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COLLEGE  OF  PHYSICIANS 
AND   SURGEONS 


Reference  Library 

Given  by 


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THE   PHYSIOLOGY   OF  THE   AMINO   ACIDS 


Courtesy  of  Prof  .  L.  B.  Mendel 


THE  PHYSIOLOGY  OF  THE 
AMINO  ACIDS 


By 
FRANK  P.  UNDERHILL,   Ph.D. 

Profeaaor  of  Pathological  Chemistry,  Yale  University 


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New  Haven:    Yale  University  Press 

London  :    Humphrey  Milford 

Oxford  University  Press 

MDCCCCXV 


and  B  show  the  contrast  between  two  rats  of  the  same 
age,  one  of  which,  B,  has  been  stunted.  The  lower  two  pictures 
afford  a  comparison  between  two  rats  of  the  same  weight,  but 
widely  differing  in  age.  The  older,  stunted  rat,  B,  has  not  lost 
the  characteristic  proportions  of  the  younger  animal,  C. 


THE  PHYSIOLOGY  OF  THE 
AMINO  ACIDS 


By 
FRANK  P.  UNDERHILL,  Ph.D. 

Professor  of  Pathological  Chemistry,  Yale  University 


New  Haven:   Yale  Univehsity  Press 

London:    Humphrey  Milford 

Oxford  University  Press 

MDCCCCXV 


COPYRXGHT,  1915 
BY 

Yale  University  Press 


First  printed,  November,  1915 
Reprinted,  June,  1916 


PREFACE 

During  the  past  few  years  the  physiology  of  the 
amino  acids  has  been  subjected  to  much  experimenta- 
tion with  the  result  that  these  protein  cleavage  prod- 
ucts have  assumed  an  ever  increasing  importance  in 
the  problems  associated  with  nitrogenous  metabolism. 
Owing  largely  to  our  too  recent  appreciation  of  the 
significance  of  these  substances  in  metabolic  processes 
there  exists  at  present  no  compilation  which  fur- 
nishes an  adequate  conception  of  the  roles  which  may 
be  played  by  the  amino  acids.  It  has  been,  therefore, 
the  aim  of  the  writer  to  gather  together  in  one  place 
the  results  which  have  thus  far  been  obtained  in  the 
field  of  the  biochemistry  of  the  amino  acids,  thus 
affording  the  busy  practitioner,  and  others  whose 
resources  for  consulting  original  communications  are 
limited,  an  opportunity  of  gaining  a  knowledge  of  the 
present-day  problems  in  this  field  of  nutrition.  In  the 
accomplishment  of  this  purpose  the  writer  has  made 
no  effort  to  include  all  the  details  or  all  the  literature 
available  upon  a  given  topic,  but  has  sought  rather  to 
indicate  leading  lines  of  thought.  At  the  end  of  each 
chapter  are  given  references  in  which  all  the  impor- 
tant literature  upon  the  topic  discussed  is  cited. 

It  is  assumed  that  the  reader  is  familiar  with  the 
fundamental  principles  of  metabolism,  hence,  in  gen- 
eral, these  have  been  omitted. 

The  author  is  deeply  indebted  to  Professor  Lafay- 
ette B.  Mendel  for  suggestions,  criticisms  of  the 
manuscript  and  for  some  of  the  plates  presented. 


CONTENTS 

Chapter  I.  page 

The  Proteins  and  their  Derivatives,  the  Amino 
Acids       .......         1 

Chapter  11. 

Digestion,  and  Bacterial  Activity  in  Relation 
to  the  Amino  Acids  .....       28 

Chapter  III. 

The    Absorption    of    Proteins    and    Amino 
Acids 46 

Chapter  IV. 

In  What  Form  Does  Ingested  Protein  Enter 
the  Circulation?       .  .  .  .         .58 

Chapter  V. 

Theories  of  Protein  Metabolism  ...  81 
Chapter  VI. 

The  Further  Fate  of  Amino  Acids  .         .       99 

Chapter  VII. 

The  Amino  Acids  in  Relation  to  the  Specific 
Dynamic  Action  of  Proteins     .         .         .     120 

Chapter  VIIL 
The  Amino  Acids  and  Simpler  Nitrogenous 
Compounds  as  Foodstuffs         .         .         .     126 

Chapter  IX. 

The  Specific  Role  of  Amino  Acids  in  Nutrition 
and  Growth     .  .  .  .  .  .136 

Index  .......     159 


t 


LIST  OF   ILLUSTRATIONS 
Plate.     Photograph  of  mice     .         .  Frontispiece 

PAGE 

Figure  1.  Survival  periods  of  mice  on  diet  of 
zein  and  tyrosine  and  zein  and 
tryptophane        ....     140 

Figure     2.     Growth  curve  of  normal  white  rats     144 

Figure     3.     Growth  curve  with  casein  and  milk 

diets  .....     144 

Figure    4.     Growth  curve  with  milk  diet  .  .     145 

Figure     5.     Maintenance  on  casein  and  growth 

after  addition  of  protein-free  milk     146 

Figure  6.  Resumption  of  growth  after  addi- 
tion of  protein-free  milk  to  casein 
diet 147 

Figure     ^.     Failure  of  growth  on  gliadin  plus 

protein- free  milk         .  .  .     149 

Figure     8.     Recovery  of  the  capacity  to  grow 

after  a  period  of  stunting  .  .     150 

Figure     9.     Maintenance    and    fertility    on    a 

gliadin   diet        .  .  .  .151 

Figure  10.     Indispensability      of      lysine      for 

growth 153 

Figure  11.  Failure  of  zein  to  maintain  or  pro- 
mote growth     ....     154 

Figure  12.     Indispensability  of  tryptophane  for 

maintenance  in  nutrition     .  .     156 

Figure  13.     Growth  on  diets  containing  zein  + 

tryptophane  -f-  lysine  .         .157 


THE  PHYSIOLOGY  OF  THE  AMINO  ACIDS 


CHAPTER  I 

THE  PROTEINS  AND  THEIR  DERIVATIVES— 
THE  AMINO  ACIDS 

The  Proteins 

The  presence  of  nitrogen  as  a  fundamental  con- 
stituent of  protoplasm  attests  the  supreme  importance 
of  this  element  for  the  construction  of  living  matter 
and  the  continued  existence  of  organized  life.  It  is 
well  recognized,  however,  that  all  forms  of  nitrogen 
are  not  equally  available  for  the  maintenance  of 
physiological  rhythm.  In  support  of  this  may  be  cited 
the  fact  that  although  the  animal  organism  is  con- 
tinually surrounded  by  an  atmosphere  rich  in  nitrogen, 
little  or  none  of  this  nitrogen  can  be  utilized  by  the 
body  for  nutritional  purposes.  The  organism  pos- 
sesses discriminating  powers  and  demands  nitrogen 
in  a  specific  form,  namely,  such  as  that  peculiar  to 
protein  and  its  derivatives.  Protein  material  con- 
stitutes therefore  an  essential  foodstuff  and  without 
it  life  would  be  impossible  for  any  considerable  period 
of  time.  "It  is  the  chemical  nucleus  or  pivot  around 
which  revolves  a  multitude  of  reactions  characteristic 
of  biological  phenomena." 


2  THE  AMINO  ACIDS 

Viewed  from  the  chemical  standpoint  protein  is 
seen  as  a  huge  molecule,  complex  in  structure,  labile 
in  character  and  therefore  prone  to  chemical  change. 
So  large  and  intricate  is  the  make-up  of  the  molecule 
that  chemists  for  generations  have  been  baffled  in 
their  attempts  to  gain  any  adequate  conception  of  its 
nature.  At  the  present  stage  of  our  knowledge  it  is 
impossible  to  form  any  satisfactory  definition  of  a 
protein  based  either  on  its  chemical  or  physiological 
properties.  In  general,  proteins  contain  about  15  to 
19  per  cent  of  nitrogen,  52  per  cent  of  carbon,  7  per 
cent  of  hydrogen,  23  per  cent  of  oxygen  and  0.5-2.0 
per  cent  of  sulphur.  Some  also  contain  phosphorus 
or  iron.  They  act  like  amphoteric  electrolytes,  that 
is,  they  are  capable  of  forming  salts  with  both  acids 
and  bases.  Proteins  belong  to  that  class  of  substances 
known  as  colloids  and  as  such  do  not  possess  the 
power  to  pass  through  animal  or  vegetable  mem- 
branes. In  a  manner  similar  to  colloids  they  may  be 
separated  from  their  solutions  by  suitable  treatment 
with  salts,  such  as  sodium  chloride,  ammonium  sul- 
phate, etc.  By  a  process  known  as  "coagulation," 
which  may  be  induced  by  the  action  of  heat  or  the 
long  continued  influence  of  alcohol  the  proteins  lose 
their  colloidal  characteristics  which  cannot  be  restored. 

Many  proteins  are  capable  of  crystallization  and 
indeed  may  occur  in  nature  in  crystalline  form.  It 
has  been  found  possible  also  to  cause  some  to  crys- 
tallize although  their  presence  in  nature  as  crystals 
is  unknown.     Some  doubt  has  been  cast  upon  the 


THE  PROTEINS  3 

probability  of  proteins,  as  we  differentiate  them  at 
present,  being  chemical  units,  but  since  many  of  the 
crystalline  plant  proteins  show  a  constancy  of  proper- 
ties and  ultimate  composition  there  is  little  reason 
for  the  assumption  that  these  at  least  are  mixtures  of 
two  or  more  individuals. 

Concerning  the  size  of  the  protein  molecule  some 
idea  may  be  gained  when  it  is  recalled  that  the  molec- 
ular weight  has  been  calculated  to  be  approximately 
15,000. 

The  proteins  possess  the  property  of  turning  the 
plane  of  polarized  light  to  the  left,  the  degree  of  rota- 
tion for  an  individual  protein  varying  with  the  solvent 
employed. 


Classification  of  Proteins 

At  present  proteins  are  classified  according  to  their 
physical  properties,  as,  for  example,  their  solubility 
in  pure  water,  weak  salt  solutions  and  dilute  acids  and 
alkalies.  It  is  well  recognized  that  such  a  classifi- 
cation is  far  from  ideal,  but  it  is  the  most  satisfactory 
plan  that  has  been  offered.  When  more  complete 
knowledge  is  gained  concerning  the  chemical  make-up 
of  the  protein  molecule  a  classification  will  undoubt- 
edly be  framed  which  will  be  based  upon  the  presence 
or  proportion  of  certain  chemical  groups  in  the  differ- 
ent proteins. 

All   albuminous   substances   may   be   divided   into 


4  THE  AMINO  ACIDS 

three  large  groups,  namely,  the  Simple  Proteins,  the 
Conjugated  Proteins  and  the  Derived  Proteins, 
Simple  Proteins  may  be  defined  as  substances  which 
yield  only  a-amino  acids  or  their  derivatives  on  hydrol- 
ysis. Conjugated  Proteins  are  substances  which  con- 
tain the  protein  molecule  united  to  some  other  mole- 
cule or  molecules  otherwise  than  as  a  salt.  As  their 
name  implies,  the  Derived  Proteins  are  substances  that 
have  been  formed  from  naturally  occurring  proteins. 
The  various  sub-divisions  of  these  large  groups,  as 
adopted  by  the  American  Physiological  Society  and 
the  American  Society  of  Biological  Chemists,  follow: 


Simple  Proteins 


Conjugated  Proteins 


1.  Albumins.  1.  Nucleoproteins. 

2.  Globulins.  2.  Glucoproteins. 

3.  Glutelins.  3.  Phosphoproteins. 

4.  Alcohol-Soluble  Proteins.  4.  Hemoglobins. 

5.  Albuminoids.  5.  Lecithoproteins. 

6.  Histones. 

7.  Protamines. 


Derived  Proteins 


A.     Primary 
Protein  Derivatives. 

1.  Proteans. 

2.  Metaproteins. 

3.  Coagulated  Proteins. 


B.     Secondary 
Protein  Derivatives. 

1.  Proteoses. 

2.  Peptones. 

3.  Peptides, 


THE  PROTEINS  5 

Occurrence   and    Characteristics   of   Different 
Classes  of  Proteins 

A.  Simple  Proteins 

Albumins  are  simple  proteins  that  are  soluble  in 
pure  water  and  are  coagulable  by  heat.  Globulins, 
on  the  other  hand,  are  insoluble  in  pure  water  but 
are  readily  soluble  in  dilute  salt  solutions.  Albumins 
and  globulins  are  generally  found  together  in  nature 
occurring,  for  example,  in  large  quantity  in  the  blood 
serum,  white  of  egg,  in  the  substance  of  cells  in  gen- 
eral, and  in  various  seeds.  Egg  white  may  be  divided 
into  two  parts  by  dialysis  against  distilled  water — 
the  globulin  being  precipitated  owing  to  the  diffusion 
of  the  salts  from  the  solution  which  originally  were 
present  in  quantity  sufficient  to  hold  the  globulin  in 
solution. 

Glutelins  are  simple  proteins  insoluble  in  all  neu- 
tral solvents  but  easily  soluble  in  very  dilute  acids  and 
alkalies.  Alcohol-Soluble  Proteins  are  simple  proteins 
readily  soluble  in  relatively  strong  alcohol  (70  to  80 
per  cent),  but  are  insoluble  in  water,  absolute  alco- 
hol and  other  neutral  solvents.  These  two  groups  of 
proteins  occur  together  as  constituents  of  the  cereal 
grains.  Glutenin  and  Gliadin,  respectively,  from 
wheat,  are  the  best  known  examples  of  these  two 
groups.  They  constitute  the  gluten  of  flour.  The 
elasticity  and  strength  of  the  gluten,  and  therefore  the 


6  THE  AMINO  ACIDS 

baking  qualities  of  a  flour  are  influenced  by  the  pro- 
portions of  glutenin  and  gliadin. 

Albuminoids  may  be  defined  as  simple  proteins 
which  possess  essentially  the  same  chemical  compo- 
sition as  the  other  proteins,  but  are  characterized  by 
great  insolubility  in  all  neutral  solvents.  Examples 
of  this  group  may  be  found  as  the  organic  basis  of 
bone  (ossein),  of  tendon  (collagen  and  its  hydration 
product,  gelatin),  of  ligament  (elastin)  and  of  nails, 
hairs,  horns,  hoofs,  and  feathers  (keratins). 

Histories  are  basic  proteins  which  may  be  looked 
upon  as  standing  between  protamines  and  the  more 
complex  proteins.  They  are  precipitated  by  other 
proteins  and  yield  a  coagulum  on  heating  which  is 
readily  soluble  in  very  dilute  acids.  The  histones  are 
soluble  in  water  but  insoluble  in  ammonia.  They  have 
been  isolated  from  varied  sources,  as  glohin  from 
hemoglobin,  scombron  from  spermatozoa  of  the  mack- 
erel, gaduhiston  from  the  codfish  and  arbacin  from 
the  sea-urchin. 

Protomines  are  the  simplest  natural  proteins.  They 
are  soluble  in  water,  are  not  coagulable  by  heat,  have 
the  property  of  precipitating  other  proteins  from  their 
solutions,  are  strongly  basic  and  form  stable  salts  with 
strong  mineral  acids.  Examples  of  protamines  are 
salmin  (from  the  spermatozoa  of  the  salmon),  sturin 
(from  the  sturgeon),  clupein  (from  the  herring),  and 
scombin  (from  the  mackerel). 


THE  PROTEINS  7 

B.  Conjugated  Proteins 

I 

Nucleoproteins  are  compounds  of  one  or  more 
protein  molecules  united  with  nucleic  acid.  The 
nucleoproteins,  as  their  name  implies,  are  the  proteins 
of  cell  nuclei  and  give  to  the  latter  their  character. 
The  nucleoproteins  are  therefore  found  in  largest 
quantity  wherever  cellular  material  is  abundant,  as 
in  glandular  tissues  and  organs.  By  artificial  hydroly- 
sis or  during  treatment  in  the  alimentary  tract  a  nucleo- 
protein  is  decomposed  into  protein  and  nucleic  acid. 
Nucleic  acid,  of  which  there  are  several  types,  may 
be  made  to  yield  a  series  of  well-defined  compounds, 
the  purine  bases  (xanthine,  hypoxanthine,  adenine  and 
guanine),  the  pyrimidine  bases  (uracile,  cytosine  and 
thymine),  a  carbohydrate  group  (pentose  or  hexose) 
and  phosphoric  acid. 

Glucoproteins  are  compounds  of  the  protein  mole- 
cule with  a  substance  or  substances  containing  a  car- 
bohydrate group  other  than  a  nucleic  acid.  Particu- 
larly rich  in  glucoproteins  are  the  mucus-yielding 
portions  of  tissues.  They  serve  also  as  a  cement  sub- 
stance in  holding  together  the  fibers  in  tendons  and 
ligaments.  An  amino-sugar,  glucosamine,  has  been 
isolated  from  some  of  the  glucoproteins  and  it  is  gen- 
erally regarded  as  constituting  the  carbohydrate  radicle 
of  these  conjugated  proteins. 

Phosphoproteins  are  compounds  of  the  protein 
molecule  with  some,  as  yet  undefined,  phosphorus- 
containing,  group  other  than  a  nucleic  acid  or  lecithin. 


8  THE  AMINO  ACIDS 

Conspicuous  foods  containing  phosphoproteins  are 
milk  with  its  caseinogen  and  tgg  yolk  with  its  vitellin. 
A  trace  of  iron  is  also  evident  in  these  proteins  and 
although  it  is  possibly  present  as  an  impurity  there 
is  no  evidence  that  it  does  not  exist  in  combination 
with  the  protein. 

Hemoglobins  are  compounds  of  the  protein  mole- 
cule with  hematin  or  some  similar  substance.  The 
coloring  matter  of  the  blood  is  hemoglobin  which 
acts  as  oxygen  carrier  for  the  tissues  and  is  charac- 
terized by  holding  iron  as  a  constituent  part  in  organic 
combination.  Globin  is  the  protein  portion  of  hemo- 
globin. In  certain  of  the  lower  animal  forms  copper 
enters  into  combination  with  protein  forming  hsemo- 
cyanin  imparting  a  blue  color  to  the  blood. 

Lecitho proteins  are  compounds  of  the  protein  mole- 
cule with  lecithins.  Lecithins  are  complexes  charac- 
terized by  yielding  glycerol,  phosphoric  acid,  fatty 
acid  radicles,  and  a  nitrogenous  base,  choline.  The 
lecithins  are  present  in  all  plant  and  animal  cells  but 
are  especially  abundant  in  the  nervous  tissues.  They 
belong  to  the  group  of  essential  cell  constituents. 

C.  Derived  Proteins 

Certain  of  the  native  soluble  proteins  upon  con- 
tinued contact  with  water,  or  the  influence  of  enzymes 
or  acid  change  their  character  and  become  insoluble. 
Such  insoluble  substances  are  called  proteans.  After 
repeated  reprecipitation  globulins  may  become  insolu- 


THE  PROTEINS  9 

ble,  that  is,  they  are  changed  to  proteans,  and  it  is 
believed  by  some  protein  investigators  that  nearly 
every  protein  may  assume  a  protean  state. 

The  metaproteins  may  be  formed  from  simple 
protein  by  the  action  of  acids  and  alkalies.  In  this 
instance,  however,  the  change  is  undoubtedly  more 
profound  than  in  the  case  of  the  proteans.  Formerly, 
metaproteins  were  termed  albuminates,  that  formed 
by  acid  being  called  acid  albuminate,  that  from  the 
action  of  alkali  being  designated  alkali  albuminate. 
These  substances  are  insoluble  in  neutral  fluids  but 
are  readily  soluble  in  an  excess  of  acid  or  alkali.  The 
metaproteins  are  of  interest  when  it  is  recalled  that 
the  acid  metaprotein  arises  as  the  first  step  in  gastric 
digestion  of  protein  and  that  likewise  alkali  meta- 
protein may  be  formed  during  pancreatic  digestion. 

The  coagulated  proteins  can  be  produced  from 
simple  proteins  by  the  long  continued  action  of  alco- 
hol, stirring  or  shaking  of  their  solutions,  or  by  the 
influence  of  heat.  In  one  instance,  namely,  the  trans- 
formation of  fibrinogen  into  fibrin  in  shed  blood,  the 
process  has  long  been  assumed  to  be  induced  by  an 
enzyme.  More  recent  work,  however,  tends  to  show 
that  enzyme  action  is  not  concerned  in  the  reaction. 

The  class  of  derived  proteins  called  Secondary 
Protein  Derivatives  represent  a  more  profound  change 
from  simple  proteins  than  is  true  for  the  proteans, 
metaproteins  and  coagulated  proteins  which  are 
grouped  together  as  Primary  Protein  Derivatives. 
Of   the  secondary  protein   derivatives  the  proteoses 


10  THE  AMINO  ACIDS 

and  peptones  are  characterized  chiefly  by  their  greater 
solubiUty  and  by  the  fact  that,  unUke  most  other  pro- 
/  teins,  they  are  diffusible  through  suitable  membranes. 
They  represent  stages  in  gastric,  pancreatic  and  bac- 
terial digestions  of  protein  and  the  peptones  are 
regarded  as  products  of  greater  cleavage  than  the 
proteoses.  There  are  several  proteoses,  as  protopro- 
teose,  heteroproteose  and  deuteroproteose  and  prob- 
ably there  may  be  several  types  of  peptones.  The 
proteoses  are  distinguished  from  the  peptones  prin- 
cipally in  being  precipitated  from  solutions  by.  satura- 
tion with  ammonium  or  zinc  sulphate. 

The  peptides  are  "definitely  characterized  combina- 
tions of  two  or  more  amino  acids,  the  carboxyl 
(COOH)  group  of  one  being  united  with  the  amino 
(NH2)  group  of  the  other  with  the  elimination  of  a 
molecule  of  water."  For  example,  if  two  molecules 
of  glycocoU  (glycine) — amino-acetic  acid — are  con- 
densed, a  peptide,  glycyl-glycine,  will  result.    Thus — 


NHH  NH, 

CH«.CO.NH 

CH,.COOH 


CH,.CO 


OH 


NH 


H 


CH2.COOH  glycyl-glycine. 

The  peptides  are  designated  di-tri-tetra-peptides,  etc., 
according  to  the  number  of  amino  acids  in  combina- 
tion.   The  name  polypeptides  is  also  applied  to  these 


THE  PROTEINS  11 

substances.  It  is  usually  accepted  at  the  present  time 
that  the  peptones  are  relatively  simple  polypeptides, 
the  line  of  demarcation  between  a  simple  peptone  and 
a  complex  peptide  not  being  well  defined. 

The  Amino  Acids 

For  nearly  a  century  chemists  have  been  seeking 
to  establish  the  composition  and  structure  of  the  pro- 
tein molecule.  Progress,  which  was  slow  and  irregular 
in  the  earlier  decades  of  this  period,  has  taken  rapid 
strides  in  the  last  twenty  years,  more  intimate  knowl- 
edge of  the  problem  being  gained  during  this  inter- 
val than  in  all  previous  time.  The  investigation  has 
been  pursued  in  three  directions — first  the  demolition 
of  the  molecule  and  the  subsequent  identification  of 
the  resulting  fragments;  second,  the  determination  of 
the  quantitative  relationships  of  these  fragments ;  and 
finally,  attempts  to  unite  the  disintegration  products 
in  such  a  manner  as  to  reproduce  the  original  molecule. 

After  a  considerable  period  of  investigation  it  was 
established  that,  although  the  protein  molecule  may 
yield  different  types  of  substances  according  to  the 
character  of  the  means  employed  for  disrupting  it 
thus  indicating  a  variety  of  possible  lines  of  cleavage, 
hydrolysis  furnishes  the  most  promising  types  of 
units.  Latterly,  this  type  of  chemical  reaction  has 
been  employed  exclusively  and  it  has  yielded  the 
important  information  now  available  concerning  the 
nature  of  the  protein  decomposition  products.     Each 


12  THE  AMINO  ACIDS 

protein  investigated  by  this  method  was  found  to 
yield  relatively  large  molecules,  such  as  proteoses  and 
peptones,  and  on  further  disintegration  a  series  of 
comparatively  simple  nitrogenous  substances  of  low 
molecular  weight  which  belong  to  a  definite  group  of 
chemical  compounds — namely,  the  amino  acids.  An 
amino  acid  may  be  regarded  as  an  organic  acid  in 
which  one  hydrogen  is  replaced  by  the  amino  group 
(NH2),  or  viewed  from  another  standpoint,  an 
amino  acid  may  be  considered  as  a  substituted  am- 
monia, one  hydrogen  of  ammonia,  NH3,  being 
replaced  by  an  organic  acid-  A  description  of  the 
amino  acids  yielded  by  proteins  follows. 

...  ^  NH2 

Glycocoll  or  glycine,  ammo-acetic  acid.     CH2.  <[  p^w^tt 

is  the  simplest  of  the  products  obtained  from  pro- 
tein by  hydrolytic  cleavage  and  it  was  also  the  first  to  be 
discovered.  Its  separation  dates  back  to  1820  in  which 
year  Braconnot  obtained  the  substance  by  boiling  gelatin  with 
sulphuric  acid,  and  because  of  its  sweet  taste  called  it  sugar 
of  gelatin.  About  twenty-five  years  later  Dessaignes  isolated 
it  after  a  hydrolysis  of  hippuric  acid.  It  was  shown  by 
Strecher  in  1848  that  glycocholic  acid,  then  called  cholic  acid, 
consists  of  a  combination  of  cholalic  acid  and  glycocoll,  and 
in  consequence  of  its  being  a  constituent  of  a  bile  acid,  glyco- 
coll assumed  a  position  of  some  physiological  importance.  Its 
presence  in  various  types  of  albuminoids,  such  as  elastin,  etc., 
was  later  demonstrated  and  finally  it  was  shown  to  be  a 
decomposition  product  of  globulin.  Glycocoll  is  not  present 
in  all  proteins  for  albumin,  casein,  and  hemoglobin  fail  to 
yield  it,  and  from  the  vegetable  proteins  it  is  obtained  in 
small  quantities  only.  On  the  other  hand,  albuminoids  are 
particularly  rich  in  glycocoll.     In  an  extract  of  the  mollusc 


THE  PROTEINS  13 

Pecten  irradians  Chittenden  found  glycocoll  in  a  free  state; 
and  it  has  been  reported  as  occurring  in  the  urine  under  vari- 
ous pathological  conditions.  After  administration  of  benzoic 
acid  to  man  and  animals  hippuric  acid  (benzoyl-glycocoll)  is 
found  in  the  urine — thus  demonstrating  a  synthesis  of  hip- 
puric acid  from  benzoic  acid  and  glycocoll. 

NH2 
Alanine — a- amino-propionic  acid,      CHs.CH  «<  p^^^^tt 

was  prepared  synthetically  previous  to  its  isolation 

from    among   the   protein    decomposition    products   and   was 

named  by  its  discoverer,  Strecher.     Alanine  has  been  shown 

to  be  a  constant  decomposition  product  of  proteins. 

CH3  NH2 

Valine — a-amino-isovalerianic  acid.  ">  CH .  CH  <" 

CH3  XOOH 

In  1856  V.  Gorup-Besanez  Isolated  a  substance  having  the 
formula  C5H11NO2  from  pancreas  and  because  it  possessed 
properties  similar  to  leucine  he  looked  upon  it  as  a  homologue 
of  leucine  and  called  it  butalanine.  Although  a  similar  sub- 
stance was  isolated  from  certain  seedlings  by  Schulze  and 
Barbieri,  and  from  the  protamine,  clupeine,  by  Kossel,  it  was 
not  until  1906  that  its  identity  was  established  by  Fischer 
who  gave  it  the  name  of  valine.  Valine  is  obtained  from  most 
proteins. 

Leucine,     a-amino-isobutylacetic  acid. 

CH3  NH2 

>CH.CH2.CH<^^^  ^ 
CHs  COOH 

Leucine  was  described  by  Proust  in  1818  and  was  called 
oxide-caseux.  Braconnot  in  1820  obtained  a  substance  from  a 
hydrolysis  of  meat  which  on  account  of  its  glistening  white 
appearance  he  called  leucine.  Liebig  regarded  it  as  one  of  the 
constituents  of  the  protein  molecule  and  this  was  later  proved 
to  be  correct.  Leucine  is  also  a  constituent  of  many  organs 
and  tissues  occurring  in  the  free  state.    It  is  yielded  by  both 


14  THE  AMINO  ACIDS 

animal  and  vegetable  proteins  and  with  the  possible  exception 
,  of  arginine  is  the  most  widely  distributed  amino  acid  found  as 
a  protein  cleavage  product.    Leucine  has  been  found  also  in 
the  urine  under  pathological  conditions. 
Isoleucine,     a-amino-/3-ethyl-propionic  acid. 

CHs  NH2 

C2H5>^^-^^<COOH 

This  amino  acid  was  not  described  as  a  protein  constituent 
until  1903  when  it  was  isolated  as  a  decomposition  product  of 
fibrin  and  other  proteins  by  F.  Ehrlich. 

Norleucine.  o-amino-normal-caproic  acid.  CH3.CH2. 
CH2.CH2.CH.NH2.COOH.  From  the  leucine  fraction  of 
the  decomposition  of  the  proteins  of  nervous  tissue  this  amino 
acid  has  recently  been  isolated  by  Abderhalden  and  Weil.  It 
is  probable  that  other  proteins  may  yield  it  also. 

Phenylalanine,    jS-phenyl-a-amino-propionic  acid. 

NH2 

Although  it  had  been  recognized  for  many  years  that  a 
substance  having  the  composition  of  C9H11NO2  could  be  ob- 
tained by  cleavage  of  both  animal  and  vegetable  proteins,  it 
was  Fischer  who  first  proved  the  presence  of  phenylalanine  as 
a  protein  derivative.  In  those  proteins  lacking  tyrosine,  as 
\/  gelatin,  for  example,  the  aromatic  ring  is  supplied  by  phenyla- 
lanine. 

Tyrosine.    /S-para-oxyphenyl-a-amino-propionic  acid. 

NH2 
HO.C6H6.CH2.CH<^^^„ 
COOH 

In  1846  Liebig  isolated  from  a  decomposition  of  cheese  a 
substance  possessing  the  property  of  crystallizing  in  silky 
needles.  He  named  it  tyrosine.  Since  then  tyrosine  has  been 
regarded  as  a  protein  cleavage  product.    It  was  not  until  18S2, 


THE  PROTEINS  15 

however,  that  the  structure  of  tyrosine  was  positively  deter- 
mined. Tyrosine  is  absent  from  the  gelatine  molecule.  In 
acute  yellow  atrophy  of  the  liver  and  in  phosphorus  poisoning 
it  is  claimed  that  tyrosine  may  be  present  as  a  urinary  con- 
stituent. 
Serine  /S-hydroxy-a-amino-propionic  acid. 

NHa 
Ca(OH).CH<^^^^ 

Cramer  found  serine  among  the  decomposition  products  of 
sericin  (silk  gelatin),  and  it  was  not  obtained  again  until  1902 
when  Fischer  isolated  it  from  various  proteins  as  a  result  of 
hydrolysis.    He  also  definitely  established  its  structure. 

Cystine,  di-cysteine  or  di-jS-thio-a-amino-propionic  acid. 
HOOC.CH.NH2.CH2.S— S.CH2.CH.NH2.COOH. 

Cystine  has  been  known  since  1810  having  been  first  de- 
scribed by  WoUaston  who  separated  it  from  a  urinary  calculus 
and  called  it  cystic  oxide.  From  that  period,  although  cystine 
was  repeatedly  isolated  from  various  organs  of  the  body,  as 
the  liver  and  kidney,  its  presence  as  a  regular  decomposition 
product  of  protein  was  not  established  until  1899  when 
K.  A.  H.  Morner  ohtained  it  by  a  hydrolysis  of  horn.  Bau- 
mann  demonstrated  the  relationship  of  cysteine  to  cystine  and 
thus  revealed  the  structure  of  cystine.  Cysteine  and  cystine 
bear  the  same  relation  to  one  another  as  does  a  mercaptan  to 
a  disulphide,  thus, 

CH2.SH  CH2 S  —  S CHj 

I  I  I 

CH.NH2  CH.NH2  CH.NHa 

I  I  I 

COOH  COOH  COOH 

cysteine  cystine 

Cystine  is  of  considerable  importance  in  metabolism  inasmuch 
as  it  is  the  only  known  sulphur-containing  amino  acid  in  the 
protein  molecule. 


16  THE  AMINO  ACIDS 

Aspartic  Acid — ^Amino-succinic  acid. 

CH2.COOH 

I 
CH.NH2.COOH 

Asparagine,  the  amide  of  aspartic  acid,  has  been  known  since 
1806,  having  been  isolated  from  asparagus  juice  by  Robiquet 
and  Vanquelin.  Upon  boiling  asparagine  with  lead  hydroxide 
Plisson  in  1827  obtained  aspartic  acid.  In  1868  aspartic  acid 
was  shown  by  Ritthausen  to  be  present  as  a  product  of  hydro- 
lytic  cleavage  of  vegetable  proteins.  In  a  similar  manner 
Kreussler  obtained  it  upon  hydrolysis  of  animal  proteins  and 
in  1874  it  was  isolated  by  Radziejeioski  and  Salkowski  from 
a  tryptic  digestion  of  protein.  Its  structure  was  established  in 
1887. 

Glutamic  Acid  (Glutaminic  Acid)   a-amino-glutaric  acid. 

CH2 

I 
CH2.COOH 

( 
CH.NH2.COOH 

Although  glutamic  acid  was  first  separated  from  a  hydrol- 
ysis of  wheat  gluten  in  1866  by  Ritthausen  its  structure  was 
not  shown  until  1890.  Ritthausen  demonstrated  that  it  was  an 
amino  acid  and  from  this  fact  together  with  its  origin  from 
gluten  gave  it  the  name  of  glutaminic  acid.  Glutamic  acid 
was  later  shown  to  arise  from  hydrolytic  cleavage  of  proteins 
of  animal  origin  as  well  as  from  those  derived  from  the  vege- 
table kingdom. 

Lysine,     a-,  e, -diamino-caproic  acid. 

H2N.CH2.CH2.CH2.CH2.CH.NH2.COOH. 

Lysine  is  widely  distributed  as  a  protein  constituent.  It 
was  first  isolated  from  casein  by  Drechsel  in  1889.    Ellinger 


THE  PROTEINS  17 

first  demonstrated  its  structure  in  1900  by  obtaining  cadav- 
erine  from  it  by  putrefaction. 
Arginine.     a-amino-5-guanidine-valerianic  acid. 

NH2 

HN  =  C  —  NH.CH2.CH3.CH2.CH.NH2.COOH. 

Among  the  products  of  a  decomposition  of  casein  Drechsel 
found  a  substance  which  he  called  lysatinine.  Later,  in  1894, 
Hedin  demonstrated  that  this  product  was  in  reality  a  mix- 
ture of  lysine  and  arginine.  Arginine  had  been  obtained  pre- 
viously by  E.  Schulze  and  Steiger  from  the  seedlings  of 
various  plants.  Urea  and  ornithine  are  among  its  decomposi- 
tion products. 

HisHdine.    jS-imidazole-a-amino-propionic  acid. 

CH 

^  \ 
N  NH 

I  I 

CH=C.CH2.CH.NH2.COOH 

Histidine  was  discovered  by  Kossel  in  1896  among  the  de-' 
composition  products  of  the  protamine  of  sturgeon  testes. 
From  the  fact  that  histidine,  arginine,  and  lysine  each  contain 
six  carbon  atoms  Kossel  called  these  three  substances  the 
hexone  bases,  and  they  were  regarded  as  a  very  important 
portion  of  the  protein  molecule.  It  was  not  until  1904  when 
the  structure  of  histidine  was  shown  by  Pauly  and  Wind- 
haus  and  Knopp  that  it  was  recognized  to  belong  to  a  group 
of  compounds  entirely  different  from  that  including  arginine 
and  lysine. 

Proline,    o-pyrrolidine-carboxylic  acid, 

CH2 CH2 

CH2        CH.COOH 


18  THE  AMINO  ACIDS 

Proline  was  first  isolated  by  Fischer  from  casein.  Its  pres- 
ence in  various  other  proteins  was  soon  shown. 

Oxyproline. 

This  amino  acid  was  prepared  from  gelatin  in  1902  by 
Fischer.  Its  structure  is  not  yet  definitely  established  although 
it  undoubtedly  possesses  one  of  the  following  formulas. 

HO.CH CH2  CH2 CH.OH 

11  II 

CH2       CH.COOH      or      CH2        CH.COOH 

Tryptophane.     /3-indole-a-amino-propionic  acid. 
C.CH2.CH.NH2.COOH 


C6H4<'  XH 

It  was  shown  in  1826  by  Tiedemann  and  Gmelin  that  when 
chlorine  or  bromine  water  is  added  to  a  tryptic  digestion 
mixture  a  violet  color  is  produced.  Stadelmann  named  the 
substance  giving  this  reaction  proteinochromogen  and  Neu- 
meister  proved  that  any  severe  treatment  of  protein  would 
cause  the  production  of  this  compound  to  which  he  gave  the 
name  tryptophane.  Hopkins  and  Cole  in  1902  isolated  from 
a  tryptic  digestion  of  casein  a  substance  which  gave  all  the 
reactions  of  tryptophane,  namely,  the  violet  coloration  with 
bromine  or  chlorine,  the  Adamkiewicz  reaction,  and  the  pro- 
duction of  indole  and  skatole  as  a  result  of  putrefaction.  In 
this  manner  the  origin  of  the  substances  characteristic  of 
putrefaction  was  made  clear.  The  structure  of  tryptophane 
was  regarded  by  Nencki  as  indole  amino  acetic  acid.  Ellinger, 
however,  showed  it  to  be  an  indole  amino  propionic  acid. 

Caseinic  Acid,  or  diamino-trioxy-dodecanic  acid.  This 
compound  has  been  isolated  by  Skraup  from  casein  only.  Its 
structure  is  still  unknown. 


THE  PROTEINS 


19 


On  inspection  of  these  formulas  it  may  be  estab- 
lished that  certain  of  the  amino  acids  are  very  closely 
related;  thus,  glycocoll,  the  simplest  of  all,  by  intro- 
duction of  the  group  (CH3)  becomes  alanine.  This 
substance  possesses  interest  because  several  of  the 
amino  acids  may  be  regarded  as  alanine  derivatives. 
By  the  replacement  of  an  (OH)  group  alanine  be- 
comes serine,  or  by  substitution  of  an  (SH)  group 
alanine  is  changed  to  cysteine.  If  the  phenyl  group 
(CeHs)  is  introduced  phenylalanine  is  obtained,  and 
the  additional  substitution  of  an  (OH)  group  leads  to 
tyrosine. 


CH, 

CH5.OH            CH,.SH 

CH.NH, 

CH.T^H,            CH.NH, 

COOH 

COOH               COOH 

Alanine 

Serine                    Cysteine 

CHa.CeHg 

CHs.CeH^.OH 

CH.NH3 

CH.NH, 

COOH 

COOH 

Phenylalanine 

Tyrosine 

The  introduction  of  the  indole  or  iminazole  group 
leads  to  the  formation  of  tryptophane  or  histidine 
respectively. 


20 


THE  AMINO  ACIDS 


CH3 

CH.NH, 

I 
COOH 

Alanine 


CH, 

CH.NH, 
COOH 


CH 

/\ 
-C      CH 


CH         C      CH 

\       /\/ 
NH       CH 

Tryptophane 


CH— NH 
\ 


CH 


CH  — C N-^  • 

CH.NH, 

I 
COOH 

Histidine 

Again  valine,  leucine  and  isoleucine  are  closely  re- 
lated structurally  as  may  be  seen  from  the  formulas 
following. 


CH3       CH3 

\ch/ 

I 
CH.NH, 

I 
COOH 

Valine 


CH3       CHj 

\ch/ 

I 

CH, 
CH.NHj 

COOH 

Leucine 


CHs       C^Hb 

\ch/ 

I 

CH.NH, 

I 
COOH 

Isoleucine 


THE  PROTEINS  21 

Viewed  from  another  standpoint  the  amino  acids 
may  be  divided  into  mono-amino  acids, — ^glycocoll, 
alanine,  valine,  leucine,  isoleucine,  phenylalanine,  tyro- 
sine, serine,  aspartic  acid,  and  glutamic  acid, — each 
containing,  as  the  name  implies  a  single  amino  (NH2) 
group — diamino  acids,  containing  two  amino  groups, 
as  arginine,  and  lysine  and  finally  the  heterocyclic  com- 
pounds as  histidine,  proline,  oxyproline,  and  trypto- 
phane. 


The  Quantitative  Relationships  of  Amino  Acids 

IN  Proteins 

The  most  serious  obstacle  to  the  quantitative  estima- 
tion of  amino  acids  in  hydrolysis  mixtures  has  been 
that  of  inadequate  methods  of  separation.  By  means 
of  the  ester  method  of  E.  Fischer  this  difficulty  has 
been  obviated  in  large  measure.  In  Table  I  below  are 
presented  figures  showing  the  yield  of  individual  amino 
acids  obtained  by  various  investigators  from  repre- 
sentative simple  proteins.  The  figures  have  not  all 
been  derived  from  use  of  the  most  exact  methods  of 
isolation,  hence  it  is  probable  that  they  may  not  repre- 
sent maximal  values  or  be  strictly  correct.  Neverthe- 
less, they  are  sufficiently  suggestive  to  demonstrate  the 
distinct  differences  that  exist  between  the  simple 
proteins. 

Table  II  undoubtedly  gives  the  most  accurate  figures 
obtainable  at  present   for  the  quantitative  yield  of 


K 

to 

§ 

H 

o 

P 

Pi 

O 

i^ 

O 

Q 

fe 

W 

t^ 

'A 

o 

M 

^ 

H 

^ 

n 

o 

O  o 

CO 

Q 

« 

^ 

M 

<) 

< 

en 

o 

^ 

w 

M 

H 

S 

O 

< 

P^ 

H( 

h 

o 

w 

Hi 

tn 

PU 

W 

M 

§ 

M 

M 

CO 

H 

>^ 

w 

< 

> 

P 

H 

a 

-u 

H 

u 

!zi 

W   w 
CO 


(z)  aoiivos 


iZ)  inqiiOE: 


{Z)  ud:^oti{Q 


{z)  /99a" 


uttunqiY 

(z)  S3a 


(e)  uRBpo 

piouxtunqiY 


21UW  s.Avoo 

(I)  upsBO 

U'i9j.oj,dO'qdsoiu 


(azrepi) 
(2)  IIX9Z 


lonooiv 


puoinxv 
too  J  J  uipuBtav 

(z) 


tuoaj  uisxaoxg 
pazinBt^SifjQ 

■  (2) 
«nnqoi£) 


(S)     (T) 
tinnqoio 


muinqxv  uinaag 
(S)     (T) 


(pooia  ssaoH) 
niqoxSomsq 
uiojj  uiqoxf) 


VOCM      -tH 


VO  00  O  T-H 


O  O  i-H  o 


O  Cvj  r-t  00 


fO  (M"r00 


(tj     (T) 


O  CO 


rH  CM 
COCV) 


ooo" 


•OTh+t 


(M  0"r  VO  t^  <M 


»-i  00       O  rH^ 

CO  vo~rvor^ev) 


N        t^  CT\  VO 

I-       Ti-iD  t^ 


evi  os~r'^coTH 


CvlO  voov        vOOOtJ- 

>o  CO  o  o  o  ^^  cvj  o 


CO  VO 
CM  O 

CO'*OOCOO»-(»Hr-l'^lOINOr-5 


•Tt-CM         tH         ■* 
■  VO      •  »H  00  O  »H  O  O 


<M  iH  O  Ol^ 


coco 


00  lO 
CO  <M 


t^oo 

OCvj 


»H  ^  VOIOO 
CO  fvj  O  CMrH 


^•+ 


Ot^TcOOO 


CM  T^     lo  o  00 
■«^  rH       oot^«o 

ioco~r'Ho,-< 

CM         1-1 


OOOV         iH  VOIO 
COCM  +  vdr-ir-J 


cvioo~r 


1-1  (^ 
cot^+ 


'S-t-H  OO  CM 


»H  Tt-  i-I  +  lO  ■*  O 


a 


o).a  ca 


c 


§l»llfl2lSf|EB-2s 


•ill 

W    Oi3   g 


0 

0 

en 

to 

<J 

•O 

rt 

h 

0) 

rfl 

0 

.•0 

MS 

^^ 

,*^ 

iii 

« 

M 

0 

V 

OS 

-M 

<J 

0 

Oi 

n 

ns 

r/l 

0 

ri 

tn 

W 

c3 

<: 

< 

s 

Tf 

(U 

n 

•d 

t3 

U 

fl 

-3 

a> 

c8 

a 

^1 
0) 

(U 

0 

id 

/3 

5^ 

<i  0  fe 

^--^ 

^^ 

,,-^ 

iH 

CM 

THE  PROTEINS 


23 


amino   acids   obtained   by   hydrolysis   from   proteins 
representing  various  groups  of  these  substances. 


Table  II. 

Quantitative  Comparison  of  Amino  Acids  Obtained 
BY  Hydrolysis  from  Proteins 

(Compiled  by  T.  B.  Osborne,  1914)*    (After  Mendel) 


Casein 


Oval-   ^T    ,. 
bumin  Gliadin 


Zein 


Edestln 


LeRumiQ 
(Pea) 


GlycocoU  

Alanine 

Valine 

Leucine 

Proline 

Oxyproline 

Phenylalanine 

Glutammic  acid 

Aspartic  acid 

Serine 

Tyrosine 

Cystine 

Histidine 

Arginine 

Lysine 

Tryptophane,  about 
Ammonia 


0.00 
1.50 
7.20 
9.35 
6.70 
0.23 
3.20 
15.55 
1.39 
0.50 
4.50 
? 

2'.  50 
3.81 
5.95 
1.50 
1.61 


0.00 
2.22 
2.50 
10.71 
3.56 

? 

5.07 
9.10 
2.20 

? 
1.77 

? 

1.71 
4.91 
3.76 
present 
1.34 


0.00 
2.00 
3.34 
6.62 

13.22 

? 

2.35 

43.66 
0.58 
0.13 
1.61 
0.45 
1.49 
2.91 
0.15 
1.00 
5.22 


0.00 
13.39 

1.88 
19.55 

9.04 
? 

6.55 
26.17 

1.71 

1.02 

3.55 
? 

0'.82 
1.55 
0.00 
0.00 
3.64 


3.80 
3.60 
6.20 

14.50 

4.10 

? 

3.09 

18.74 
4.50 
0.33 
2.13 
1.00 
2.19 

14.17 
1.65 
present 
2.28 


0.38 

2.08 

? 

8'.  00 

3.22 

? 

3*.  75 
13.80 

5.30 

0.53 

1.55 
? 

2.42 
10.12 

4.29 
present 

1.99 


65,49       48.85       84.73 


88.87       82.28       57.43 


♦These  analyses  are  combinations  of  what  appear  to  be  the  best  de- 
terminations of  various  chemists. 


It  may  be  seen  from  these  tables  that  certain  pro- 
teins, as  serum  albumin  and  casein  contain  no  glyco- 
coll,  whereas  serum  globulin  contains  a  small  amount 
and  gelatin  a  large  quantity.  Alanine  presents  vari- 
able figures  but  is  usually  present.  The  same  may  be 
said  of  leucine,  phenylalanine,  proline,  and  aspartic 
acid.    Tyrosine  may  be  absent  as  in  gelatin.    Glutamic 


24  THE  AMINO  ACIDS 

acid  may  show  very  wide  variations  being  present  to 
the  extent  of  nearly  44  per  cent  in  wheat  gliadin  where- 
as gelatin  contains  less  than  1  per  cent.  Tryptophane 
may  be  absent  as  in  zein  and  gelatin.  Arginine  shows 
great  variation  being  present  in  largest  quantity  in  the 
protamines  (salmine).  On  the  other  hand,  lysine  is 
absent  in  salmine  as  well  as  in  zein.  In  the  protamine, 
salmine,  histidine  is  not  present  but  may  be  isolated 
from  all  other  examples  of  simple  proteins  shown  here. 
It  is  clear  that  in  general  the  various  proteins  are  made 
up  of  the  same  units  and  it  undoubtedly  follows  that 
the  individual  protein  characteristics  are  bestowed  by 
the  relative  proportion  of  the  units  or  by  their  absence. 
In  the  tables  given  it  will  be  observed  that  in  most 
instances  the  total  amino  acids  fall  far  short  of  the 
theoretical  yield,  a  deficit  of  40  to  50  per  cent  being  in 
order.  Previously  it  has  been  assumed  that  only  a 
portion  of  the  amino  acids  was  known.  At  present, 
however,  it  seems  very  probable  that  the  deficit  is  to 
be  explained  on  the  hypothesis  of  inadequate  methods 
of  analysis. 


Synthetic  Proof  of  the  Structure  of  Protein 

Since  the  time  of  Liebig  it  has  been  assumed  that 
the  protein  molecule  consisted  of  a  huge  complex  of 
amino  acids  linked  together  in  some  unknown  manner. 
There  are  many  possibilities  for  such  combinations 
and  certain  of  them  have  been  subjected  to  experimen- 


THE  PROTEINS  26 

tation  without,  however,  yielding  any  very  far-reaching 
conclusions.  It  remained  for  Emil  Fischer  and  his 
associates  in  1901  to  conceive  of  a  combination  which 
undoubtedly  will  ultimately  lead  to  a  clear  under- 
standing of  the  structure  of  the  protein  molecule. 
These  combinations  of  amino  acids  were  termed 
polypeptides.  Just  as  we  have  mono-,  di-,  or  tri- 
saccharides  so  there  may  be  di-,  tri-,  etc., -peptides. 
According  to  Fischer's  method  the  amino  acids  are 
linked  together  by  dehydration  of  their  hydroxy  1  and 
amino  groups,  the  carboxyl  group  of  each  acid  being 
condensed  with  the  amino  group  of  its  neighbor  in  the 
molecule,  thus 

NHH 


R.  CH.CO 

I 

NH 


OH 
H 


R    .CH.COOH 

By  continued  union  of  amino  acids  infinite  possi- 
bilities of  complexes  are  presented.  Actually  com- 
pounds containing  as  many  as  eighteen  amino  acids 
have  been  synthesized  by  Fischer  and  some  of  the 
products  obtained  have  shown  properties  similar  to 
those  of  the  native  protein. 

After  demonstration  of  the  possibility  of  forming 
protein-like  compounds  by  synthesis  Fischer  next 
attempted  to  determine  whether  similar  simple  com- 
plexes  could  be  derived  from  proteins  by  suitable  treat- 


26  THE  AMINO  ACIDS 

ment.  For  this  purpose  he  employed  a  mild  hydrolysis 
which  only  partially  broke  up  the  large  aggregates 
formed  and  he  succeeded  in  isolating  from  the  pro- 
ducts peptides  identical  with  those  made  synthetically. 
Since  then  other  investigators  have  separated  similar 
compounds.  One  of  the  best  proofs  that  proteins  are 
built  up  of  these  amino  acid  complexes  is  that,  also 
furnished  by  Fischer,  of  the  action  of  various  enzymes 
upon  the  synthetical  products.  It  was  found  that  with 
the  exception  of  pepsin  the  various  enzymes  of  the 
^  body  are  quite  capable  of  hydrolyzing  the  polypeptides 
into  amino  acids. 

Although  these  investigations  prove  beyond  doubt 
that  amino  acids  are  linked  together  in  protein  in  the 
form  of  polypeptides,  there  are  possibilities  of  other 
forms  of  combination  which  will  be  revealed  only  by 
future  research.  For  the  present  we  are  justified  in 
accepting  the  hypothesis  of  the  protein  molecule  as  a 
huge  complex  polypeptide. 


References  to  Literature 


Abderhalden:  Text  Book  of  Physiological  Chemistry.    1914. 

Ahderhalden  and  Weil:  Zeitschrift  fiir  physiologische  chemie. 
1913,  88,  p.  272.    [Norleucine.] 

Hammarsten:  Text  Book  of  Physiological  Chemistry.     1914. 

Kossel:  The  Chemical  Composition  of  the  Cell.    The  Harvey 
Lectures.    1911-1912. 


THE  PROTEINS  27 

Kossel:  The  Proteins.  Johns  Hopkins  Bulletin.  1912,  23, 
p.  65. 

Mann:  Chemistry  of  the  Proteins.    1906. 

Mendel:  Nutrition  and  Growth:  Harvey  Lectures  1914-15. 
Journal  of  the  American  Medical  Association.  1915,  64, 
p.  1539. 

Osborne:  The  Vegetable  Proteins,  1909. 

Osborne:  Chemistry  of  the  Proteins.  The  Harvey  Lectures, 
1910-1911. 

Plimmer:  Chemical  Constitution  of  the  Proteins,  1908. 

Van  Slyke:  The  Proteins.  New  York  Medical  Journal.  1912, 
August  10  and  17. 


CHAPTER  II 

DIGESTION,  AND  BACTERIAL  ACTIVITY  IN 
RELATION  TO  THE  AMINO  ACIDS 

Concerning  the  nature  of  protein  digestion  Schaefer 
in  1898  wrote :  "The  products  found  toward  the  end  of 
a  proteid  digestion  in  vitro  are  distinguished  from  the 
proteids  from  which  they  originate  by  being  slightly 
diffusible.  To  this  fact  great  importance  was  at  one 
time  attributed,  because  it  was  thought  that  only  pro- 
teids in  a  diffusible  form  were  capable  of  absorption, 
and  hence  that  peptonization  was  in  all  cases  a  neces- 
sary preliminary.  It  is  now  generally  admitted  that 
many  forms  of  native  proteid  are  capable  of  entering 
the  epithelial  cells  (of  the  intestine)  without  previous 
change  by  digestion  or  otherwise ;  and  in  those  cases  in 
which  a  proteid  is  incapable  of  direct  absorption  a 
much  less  profound  change  than  peptonization  is  suffi- 
cient to  render  it  so,  namely,  conversion  into  acid  or 
alkali  albumin."  With  regard  to  the  extent  of  amino 
acid  formation  in  digestion  Schaefer  says:  "It  is  not 
known  with  certainty  to  what  extent  amino  acids  are 
formed  from  proteids,  in  the  natural  course  of  intes- 
tinal digestion.  The  experimental  evidence  is  some- 
what conflicting,  but  the  majority  of  observers  are  of 
the  opinion  that  but  little  proteid  is  absorbed  as  leucine 
or  tyrosine,  being  nearly  all  absorbed  as  albumose  or 


DIGESTION  29 

peptone,  or  even  at  a  still  earlier  stage.  The  only  posi- 
tive evidence  as  to  the  formation  of  leucine  and 
tyrosine  in  natural  digestion,  rests  on  the  amounts 
found  in  the  intestinal  contents  during  protein  diges- 
tion." It  is  then  stated  that  in  general  the  quantities 
of  amino  acids  present  during  digestion  are  small. 

In  the  few  years  since  the  above  was  written  the 
advances  made  in  the  chemistry  of  the  proteins  and  of 
digestion  have  made  necessary  a  radical  revision  of  our 
ideas  of  the  nature  and  extent  of  the  alimentary  treat- 
ment of  protein.  No  longer  tenable  is  the  view  that 
digestion  stops  with  the  transformation  of  insoluble 
and  non-diffusible  substances  into  compounds  soluble 
and  diffusible,  nor  can  the  idea  be  accepted  of  a  dis- 
tinction between  directly  assimilable  and  non-assimi- 
lable proteins.  The  change  to  "peptone"  is  now  held  to 
be  merely  an  intermediate  stage  in  digestion,  not  the 
end,  as  was  once  assumed.  According  to  the  latest  con- 
ception of  protein  digestion  a  profound  disintegration 
occurs,  the  ultimate  products  formed  being  a  variety  of 
polypeptides  and  amino  acids.  Digestion,  in  accord- 
ance with  this  idea,  consists  in  a  series  of  hydrolytic 
cleavages  which  are  induced  through  the  agencies  of 
the  enzymes  present  in  the  gastro-enteric  tract.  The 
products  formed  by  these  enzymes  undoubtedly  are 
identical  with  those  produced  outside  the  body  by 
means  of  the  action  of  acids.  Amino  acids  therefore 
must  be  looked  upon  as  the  ultimate  nitrogenous  food- 
stuffs— it  is  to  these  substances  that  the  organism  must 
look  for  its  essential  requirement  of  nitrogen. 


30  THE  AMINO  ACIDS 

Are  Amino  Acids  Formed  during  Gastric  Digestion 

OF  Protein? 

Protein  digestion  is  initiated  in  the  stomach  through 
the  action  of  gastric  juice — the  active  constituents 
being  pepsin  and  hydrochloric  acid.  In  investigating 
the  nature  and  extent  of  gastric  digestion  three  general 
methods  have  been  employed — as  follows:  (1)  the 
stomach  tube  method,  the  only  procedure  applicable  to 
man,  whereby  the  stomach  contents  are  withdrawn  at 
intervals  after  a  meal,  (2)  animals  fed  definite  diets 
are  quickly  killed  at  varying  periods  of  time  and  the 
stomach  contents  examined ;  or  animals  are  killed  with 
the  stomach  empty,  food  introduced  and  analyzed  at 
intervals,  (3)  the  polyfistulous  method — ^fistulas  being 
inserted  in  the  stomach  and  at  various  points  in  the 
intestine,  the  food  products  being  withdrawn  through 
these  openings. 

What  products  are  formed  in  the  stomach  under  the 
influence  of  peptic  digestion?  It  is  self-evident  that 
experiments  carried  out  under  artificial  conditions,  as 
in  beakers,  can  afford  no  positive  assurance  that  the 
products  are  identical  with  those  formed  in  the  stom- 
ach. Kiihne  was  the  first  to  demonstrate  that  pepsin 
digestion  in  vitro  leads  only  to  the  formation  of  pro- 
teoses and  peptones.  Oa  the  other  hand,  numerous 
recent  investigations  have  shown  by  the  polyfistulous 
method  that  under  normal  conditions  also  proteoses  and 
peptones  represent  the  final  stages  in  gastric  digestion 
of  protein.    All  the  protein  does  not  of  necessity  reach 


DIGESTION  31 

the  peptone  stage.  Indeed,  in  general  the  process  goes 
only  as  far  as  the  proteose  stage  as  may  be  seen  from 
the  following  table  from  London.  In  these  experi- 
ments dogs  were  fed  different  types  of  proteins  and 
from  a  fistula  below  the  pylorus  the  products  were 
collected. 

Kind  of  Protein  Fed  Percentage  of  Proteoses  Found 

Egg  Albumin  72.5 

Gliadin  67.7 

Edestin  60.3 

Casein  59.1 

Gelatin  50.6 

Serum  Albumin  46.1 

London  and  his  co-workers  also  found  that  upon 
feeding  varying  quantities  of  the  same  protein  a  defi- 
nite proportion  of  proteoses  was  always  formed,  thus — 

Quantity  of  Gliadin  Fed  Percentage  of  Proteoses  Found 
in  grams 

25  80.8 

50  86.1 

75  86.5 

100  84.9 

It  is  therefore  probable  that  ingested  protein  enters 
the  duodenum  largely  in  the  form  of  proteoses  and  to 
a  smaller  extent  as  peptones^. 

By  long  continued  action  of  both  artificial  and  nor- 
mal gastric  juice  various  investigators  have  observed 
the  gradual  formation  of  amino  acids.  These  results 
obtained  from  digestive  mixtures  allowed  to  stand  for 
months  cannot  be  regarded  as  applicable  to  normal 


32  THE  AMINO  ACIDS 

stomach  digestion  which  is  at  most  a  matter  of  hours. 
They  may  be  explained  in  several  ways,  as  for  instance 
in  those  cases  where  extracts  of  the  stomach  were 
employed  amino  acids  may  arise  from  autolytic  proc- 
esses, or  perhaps  in  all  cases  from  the  action  of  hydro- 
chloric acid  alone.  Protein  is  a  labile  molecule  which 
apparently  needs  slight  inducement  to  start  on  the 
downward  path  to  its  demolition  into  amino  acids. 
The  formation  of  amino  acids  in  gastric  digestion 
under  normal  conditions  seems  hardly  probable. 
Against  such  an  idea  may  be  set  the  fact  that  pepsin- 
hydrochloric  acid  is  utterly  incapable  of  breaking 
down  artificial  polypeptides  thus  far  tested.  On  the 
other  hand,  they  are  readily  split  by  pancreatic  juice. 
It  would  seem  therefore  that  in  peptic  digestion 
neither  amino  acids  nor  relatively  simple  polypeptides 
are  normally  found  in  significant  amounts. 

Gastric  digestion,  however,  has  the  very  important 
function  of  preparing  protein  for  the  later  action  of 
trypsin  and  the  intestinal  juices.  Fischer  and  Abder- 
halden  have  shown  that  tryptic  digestion  is  much  more 
rapid  and  complete  when  protein  has  been  previously 
acted  upon  by  pepsin-hydrochloric  acid.  If  casein  is 
first  digested  with  an  artificial  gastric  juice  and  then 
subjected  to  the  influence  of  trypsin  amino  acids  like 
proline  and  phenylalanine  could  be  isolated.  Treated 
with  trypsin  alone  casein  failed  to  yield  the  free  amino 
acids ;  instead  a  corresponding  polypeptide  was  present. 

One  may  conclude  therefore  that  although  gastric 
digestion  fails  to  yield  amino  acids  directly  it  aids  in 


DIGESTION  33 

their   rapid   formation  indirectly   by   facilitating  the 
action  of  trypsin. 

Intestinal  Digestion 

Kiihne  made  the  important  discovery  that  there  is 
an  essential  difference  between  the  digestive  action  of 
trypsin  and  that  of  pepsin.  He  stated  that  the  influ- 
ence of  the  former  does  not  cease  with  the  formation 
of  peptone  but  is  carried  to  a  stage  where  crystalline 
products  appear — the  amino  acids.  As  late  as  1900, 
however,  these  substances  were  regarded  as  by-pro- 
ducts in  natural  digestion — of  little  significance  and 
formed  in  relatively  small  quantities.  At  that  time  the 
cleavage  products  recognized  were  leucine,  tyrosine, 
aspartic  acid,  glutamic  acid,  lysine,  arginine  and  histi- 
dine  and  proteinochromogen  (see  Chapter  I).  With 
the  growth  of  knowledge  concerning  protein  chemistry 
most  of  the  characteristic  amino  acids  have  since  been 
isolated  from  intestinal  contents. 

In  1906  Cohnheim  gave  a  new  meaning  to  intestinal 
digestion  by  his  discovery  of  an  enzyme  capable  of 
splitting  proteoses  and  peptones  into  simpler  products. 
Cohnheim  was  of  the  opinion  that  synthesis  of  protein 
from  peptones  occurred  in  the  intestinal  wall.  While 
endeavoring  to  determine  this  point  he  noted  that  the 
characteristic  peptone  reaction  disappeared.  Its  dis- 
appearance was  not  due  to  protein  synthesis  as  was 
early  assumed,  but  because  crystalline  decomposition 
products  were  formed  from  it.    This  chemical  trans- 


s 


34  THE  AMINO  ACIDS 

formation  was  shown  to  be  enzymatic  in  nature  and  to 
the  enzyme  Cohnheim  gave  the  name  erepsin.  Later 
investigators  showed  that  erepsin  is  quite  specific  in 
its  action — it  has  no  influence  upon  native  proteins 
with  the  exception  of  casein  and  gelatin — but  is  capable 
of  completely  transforming  proteoses  and  peptones  into 
amino  acids,  such  as  leucine,  tyrosine,  lysine,  histidine, 
and  arginine.  In  intestinal  digestion,  therefore,  two 
agencies  are  to  be  considered  in  protein  disintegration, 
namely,  trypsin  and  erepsin.  From  these  two  differ- 
ent types  of  activity  one  may  perhaps  draw  the  con- 
clusion that  there  is  a  purposeful  function  for  each. 
It  may  be  imagined  for  instance  that  trypsin  may  per- 
form a  twofold  function,  the  degradation  of  the 
protein  molecule  which  may  have  escaped  gastric  diges- 
tion to  the  proteose  or  peptone  stage,  or  completely  to 
amino  acids.  Erepsin  on  the  other  hand  is  present  to 
guarantee  that  all  complicated  structures  as  proteoses, 
peptones,  or  polypeptides  are  reduced  to  their  simplest 
terms.  It  is  apparent,  therefore,  from  the  distribution 
of  enzymes  in  the  intestinal  tract  that  there  is  a  natural 
provision  for  ingested  protein  to  be  subjected  to  a 
series  of  hydrolytic  cleavages  whereby  only  relatively 
simple  amino  acids  are  finally  present. 

Although  it  was  generally  admitted  that  protein  di- 
gestion may  proceed  to  the  stage  of  amino  acids  it  was 
exceedingly  difficult  to  prove  the  fact  when  applied 
to  the  alimentary  tract  under  normal  conditions.  The 
difficulty  was  twofold  in  nature.  In  the  first  place, 
demolition  of  the  protein  molecule  is  not  of  the  nature 


DIGESTION  35 

of  an  explosion  resulting  in  a  large  number  of  frag- 
ments scattered  about,  but  instead  it  may  be  looked 
upon  as  a  kind  of  slow  erosion  whereby  certain  pro- 
jecting pieces  are  rubbed  or  broken  off.     Secondly, 
absorption  takes  place  rapidly  and  the  erosion  products 
have  a  tendency   to   disappear   from  the  alimentary 
canal.     A  knowledge  of  the  thorough   character  of 
intestinal  digestion  has  been  made  possible  through  the 
employment  of  the  polyfistulous  method  devised  by 
London.    Animals  with  a  series  of  fistulas  along  the 
intestinal  tract  were  fed  gliadin  and  from  successive 
openings  the  enteric  contents  were  examined  for  the 
quantity  of  tyrosine  and  glutamic  acid  present.    It  was 
shown  that  in  the  duodenal  contents  0.75  gram  tyrosin 
and  2.5  grams  of  glutamic  acid  were  present,  in  the 
jejunum  were  1.1  gram  tyrosin  and  20.9  grams  of 
glutamic  acid  w^hile  the  ileum  yielded  only  a  trace  of 
tyrosin  and  33  grams  of  glutamic  acid.    Similar  experi- 
ments with  casein  and  meat  yielded  comparable  results. 
From  these  observations  it  is  quite  evident  that  the 
processes  of  digestion  in  the  intestine  are  gradual  in 
nature  but  the  rate  of  disintegration  is  much  greater 
than    obtains    in   artificial   digestion   mixtures.      The 
apparent  explanation  for  the  slower  rate  of  hydrolysis 
in  in  vitro  experiments  is  that  the  digestion  enzymes 
form  compounds  with  the  amino  acids  split  off  and  thus 
are  rendered  inactive.    This  inactivation  probably  does 
not  occur  to  any  extent  in  the  intestine  because  the 
amino  acids  do  not  accumulate  therein,  undoubtedly 
being  absorbed  almost  as  soon  as  they  are  split  off. 


36  THE  AMINO  ACIDS 

The  small  intestine,  therefore,  may  be  regarded  as 
the  seat  of  profound  protein  digestion,  the  products 
arising  being  the  amino  acids  typical  for  hydrolytic 
cleavage  of  protein.  Undoubtedly  all  digestible  pro- 
teins are  ultimately  reduced  to  the  condition  of  amino 
acids.  From  this  it  follows  according  to  present  views 
that  nitrogenous  metabolism  is  concerned  mainly  with 
the  amino  acids  and  the  transformations  which  they 
undergo. 

Intestinal  Bacteria  and  the  Amino  Acids 

In  the  early  days  of  the  history  of  protein  digestion 
great  difficulty  was  experienced  in  the  determination  of 
the  actual  products  formed  because  of  the  accompani- 
ment of  putrefaction.  This  was  especially  true  for 
tryptic  digestion  where  it  is  desirable  to  maintain  an 
alkaline  medium,  an  environment  also  favorable  for 
bacterial  growth.  Kiihne  was  the  first  to  demonstrate 
the  activity  of  trypsin  in  the  presence  of  antiseptics 
and  through  the  employment  of  antiseptic  digestion 
mixtures  a  sharp  division  line  was  soon  drawn  between 
the  products  of  tryptic  digestion  and  those  formed  by 
bacterial  agencies. 

In  general  the  products  of  putrefaction  are  identical 
whether  formed  outside  the  body  or  within.  The  type 
of  action  is  similar  to  other  kinds  of  digestion  activity. 
Indeed,  there  is  little  doubt  that  the  same  kind  of 
agencies  are  at  work  in  the  two  instances,  namely, 
enzymes.    In  the  one  case  they  are  present  in  a  secre- 


DIGESTION  37 

tion,  as  in  intestinal  juice,  in  the  other  instance  they  are 
contained  within  an  organism.  In  bacterial  digestion 
the  first  stages  of  digestion  are  very  similar  to  those 
induced  by  trypsin.  If  the  protein  is  insoluble  solution 
is  first  effected  which  is  not  a  rapid  process  as  in  the 
case  of  trypsin.  The  proteoses  and  peptones  are  next 
formed  but  are  quickly  transformed  into  lower  decom- 
position products.  Proteoses  and  peptones  are  much 
more  readily  attacked  than  are  the  native  proteins, 
which  may  not  begin  to  undergo  a  profound  change 
until  the  former  have  been  broken  up  to  smaller  mole- 
cules. 

Putrefaction  may  be  regarded  as  causing  a  different 
type  of  cleavage  than  occurs  in  ordinary  tryptic  or  in- 
testinal digestion  as  exemplified  by  the  specific  sub- 
stances produced.  It  would  appear  much  more  likely, 
however,  that  the  early  stages  of  tryptic  digestion  and 
those  induced  by  bacteria  are  identical  in  both  instances, 
amino  acids  being  the  final  products.  On  the  other 
hand,  little  or  no  putrefaction  occurs  in  the  small  intes- 
tine and  there  is  little  reason  to  assume  that  under 
ordinary  circumstances  any  unchanged  protein  or 
perhaps  even  proteoses  or  peptones  succeeds  in  pass- 
ing the  ileo-caecal  valve.  It  is  therefore  probable  that 
normally  putrefactive  bacteria  act  upon  the  amino  acids 
rather  than  upon  their  precursors,  the  complex  protein 
molecules.  It  is  even  doubted  whether  pure  solutions 
of  native  proteins  will  putrefy  directly.  Accepting  the 
hypothesis  that  it  is  the  amino  acids  which  are  con- 
cerned primarily  in  putrefactive  processes  the  forma- 


38 


THE  AMINO  ACIDS 


tion  of  the  substances  characteristic  of  putrefaction  is 
readily  understood.  The  amino  acids  which  are  espe- 
cially susceptible  to  bacterial  action  are  tyrosine  and 
tryptophane.  From  tyrosine  a  whole  series  of  com- 
pounds may  be  formed  and  are  regularly  present  as 
putrefactive  products,  as  for  example,  paroxyphenyl- 
propionic  acid  (hydro-paracumaric  acid),  and  paroxy- 
phenylacetic  acid  (also  phenylpropionic  and  phenyl- 
acetic  acids),  as  well  as  paracresol  and  phenol.  The 
relationships  are  readily  seen  from  the  following 
formulae : 


OH 


CH, 


OH 


CH, 


OH 
/\ 


CH, 


CH.NH, 


CH, 


COOH 


COOH 

Tyrosine,  or 

p.  oxy-phenyl 

a-amino-propionic 

acid 


COOH 

p.  oxy-phenyl 

propionic 

acid 


p.  oxy-phenyl 

acetic 

acid 


OH 


CH3 
p.  cresol 


OH 


Phenol 


DIGESTION 


39 


From  tryptophane  the  malodorous  bodies  indole  and 
skatole  may  be  produced,  thus  : 


C.CH2CH.NH2.COOH 
CH 


C.CH2CH2.COOH 
CH 


Indole-amino- 
propionic  acid 
Tryptophane 


C.CHs 


NB. 


Indole- 
propionic  acid 


CH 


CH 


CH 


Indole-acetic 
acid 


NH 
Indole 


In  the  explanation  of  these  changes  in  both  instances 
it  is  seen  that  the  types  of  chemical  reactions  are  iden- 
tical. First  deamination  or  splitting  off  of  ammonia, 
NH3,  occurs.  This  is  followed  by  a  cleavage  of  carbon 
dioxide,  oxidation,  and  finally  demethylation.  The 
chemical  transformations  therefore  are  quite  varied 
and  extensive. 

When  putrefaction  is  mentioned  one  invariably 
thinks  of  indole,  skatole,  the  oxy  acids,  etc.  These 
compounds,  however,  by  no  means  represent  all  of  the 
substances  actually  formed  for  a  type  of  chemical 
compound  has  been  isolated  which  is  also  peculiarly 


40 


THE  AMINO  ACIDS 


characteristic  of  putrefaction — namely,  the  amines. 
Dixon  and  Taylor  in  1907  aroused  considerable  interest 
by  the  publication  of  their  observation  that  alcoholic 
extracts  of  the  human  placenta  when  injected  intro- 
venously  caused  a  marked  rise  in  blood  pressure  and 
contractions  of  the  pregnant  uterus.  It  was  later 
shown  that  these  phenomena  failed  to  appear  in  placen- 
tal extracts  free  from  putrefaction.  Evidence  was  soon 
produced  showing  that  putrefaction  of  the  placenta 
caused  the  production  from  tyrosine  of  a  new  body, 
namely  p-oxyphenylethylamine.  This  substance  was 
isolated  from  a  pancreas  digestion  several  years  pre- 
viously by  Emerson  and  its  production  as  a  product  of 
tryptic  action  was  regarded  as  unique.  In  the  light  of 
present  knowledge  there  is  little  doubt  that  here  also 
it  was  formed  through  bacterial  agency.  This  new 
substance  is  produced  by  the  liberation  of  CO2  from 
tyrosine,  thus: 


OH 


OH 


/ 


CH, 
CH.NH, 


|COO|H 

Tyrosine 


CH, 
CH, 
NH, 

p.  oxyphenyl-ethylamlne 


DIGESTION 


41 


To  this  compound  has  been  given  the  name  tyramine. 
It  is  of  special  significance  both  from  the  chemical  and 
pharmacological  standpoints  because  of  its  resemblance 
in  both  respects  to  epinephrine. 


OH 


CH, 

I 
CH, 

NH, 

Tyramine 


OH 

lOH 

\/ 
CH.OH 

I 
CH, 

I 
NH.CH3 

Epinephrine 


Tyramine  acts  upon  the  sympathetic  nervous  system 
as  does  epinephrine.  Its  action,  however,  is  somewhat 
weaker.  Its  effects  are  produced  whether  absorbed 
from  subcutaneous  tissues  or  from  the  alimentary 
canal.  A  further  interest  attaches  to  tyramine  in  that 
it  is  one  of  the  substances  that  confers  upon  ergot  its 
characteristic  action  on  the  uterus. 

Not  only  is  tyramine  found  in  putrefaction  mixtures 
without  the  body,  but  it  has  been  isolated  from  the 
contents  of  the  large  intestine  and  it  may,  therefore, 
be  looked  upon  as  a  product  formed  regularly  in  the 
body.  On  the  other  hand  its  presence  in  the  alimentary 
canal  does  not  necessarily  imply  that  it  was  formed 


42 


THE  AMINO  ACIDS 


there  for  it  has  been  shown  quite  recently  that  it  may 
be  ingested  with  certain  food  products.  Thus  tyramine 
occurs  in  such  varieties  of  cheeses  as  the  Camembert, 
Roquefort,  Emmenthal  and  even  the  American  cheddar 
cheese  is  not  free  from  it. 

In  a  manner  similar  to  the  formation  of  tyramine 
we  may  have  amines  produced  from  other  amino  acids 
by  bacteria.  From  leucine  may  be  formed  isoamyla- 
mine,  thus : 


CH3 


CH, 


CH. 


CH. 


CH 

CH 

CH« 

CH, 

CH.NH, 

CH, 

COOH 

NH, 

Leucine 

Isoamylamine 

From  tryptophane  a  corresponding  amine  may  be 
produced,  thus : 


x\ 

C.CH2.CH.NH2.COOH 

/X 

C.CH2.CH2.NH2 

/         \ 

rNcH 

fNcH 

\/      NH 

\/      NH 

Tryptophane 

Indo 

le-ethylamine 

When  histidine  is  subjected  to  the  action  of  putre- 
factive bacteria  it  is  transformed  to   )8-iminazolylethyl- 


DIGESTION  43 

amine  or  as  it  has  been  called  histamine.    The  reaction 
occurring  follows. 


CH 

HN    ^ 

CH 
HN     ^ 

HC C 

HC C 

CH, 

CH, 

CH.NH, 

CH, 

COOiH 

NH, 

Histidine 

/S-iminazolylethyla 

This  substance  besides  possessing  an  action  upon 
the  nervous  system  is  capable  of  producing  symptoms 
identical  with  those  of  anaphylactic  shock.  Its  pres- 
ence in  the  alimentary  canal  has  also  been  demon- 
strated. 

The  diamines,  cadaverine  and  putrescine,  arise  in 
the  alimentary  through  the  action  of  bacteria.  Cadav- 
erine is  produced  in  the  following  manner,  lysine  serv- 
ing as  the  mother  substance.  Lysine,  which  has  the 
following  formula : 

CH,.CH,.CH,.CH,.CH.COOH 

NH,  NH, 

by  cleavage  of  carbon  dioxide  yields  cadaverine  which 
has  the  structure  belo\\i  mriArkw  -.^  — .^ 

^IBf^ARY  OPTUS 

MuTflVm'^'  ■ASSOCIATION 

WtUMBIA  UNIVEksn  V 


44  THE  AMINO  ACIDS 

CHj.CHa.CHa.CHa.CHj 

NHa  NH, 

Cadaverine  or  Pentamethyldiamine 

Arginine,  another  amino  acid,  is  the  mother  sub- 
stance of  putrescine.  Arginine  under  suitable  condi- 
tions yields  urea  and  ornithine,  thus : 

CHa.CHa.CHa.CH.COOH 


NH 

NH,                  ^ 

NH, 

Arginine                                    • 

NH, 

CH,.CH,.CH,.CH.COOH 

C  =  O        + 

NH,                  NH, 

NH, 

Urea 

Ornithine 

Ornithine   by    cleavage    of    carbon    dioxide    yields 
putrescine  or  tetramethyldiamine. 

CH,.CH,.CH,.CH.COOH  CH,.CH,.CH,.CH, 

NH,  NH,         -CO,         NH,  NH, 

Ornithine  Putrescine  or 

Tetramethyldiamine 


DIGESTION  45 

It  is  exceedingly  probable  that  the  purpose  of  pro- 
tein digestion  is  the  reduction  of  these  complex  mole- 
cules to  the  form  of  crystalline  products,  the  amino 
acids.  Putrefaction  is  also  concerned  with  these  sub- 
stances forming  from  them  compounds  which  may 
exert  perhaps  at  times  a  more  or  less  deleterious 
action  as,  for  example,  indole  or  skatole,  but  also  trans- 
forming the  amino  acids  into  products,  as  tyramine  or 
histamine,  which  time  may  show  to  have  distinct  physi- 
ological activities  in  keeping  normal  the  adjustment 
of  nutritional  rhythm. 

References  to  Literature 

Barger:  The  Simpler  Natural  Bases.    1914. 

Cathcart:  Physiology  of  Protein  Metabolism.     [Digestion.] 

Hammarsten:  Text  Book  of  Physiological  Chemistry.     1914. 
[Digestion.] 

London:  Handbuch  der  Biochemie.    Oppenheimer.    III.    1909. 
[Digestion.] 

Retiger:  Journal  of  Biological  Chemistry.     1915,  20,  p.  445. 
[Putrefaction  of  pure  proteins.] 

Schaefer:  Text  Book  of  Physiology.     1898.     [Digestion.] 

Underhill:  Middleton  Goldsmith  Lecture  for  1911.    Archives 
of  Internal  Medicine,     1911,  8,  p.  356.     [Putrefaction.] 

Winterstein  and  Trier:  Die  Alkaloide.    1910. 


CHAPTER  III 

THE  ABSORPTION   OF  PROTEINS  AND 
AMINO  ACIDS 

The  views  which  have  been  held  from  time  to  time 
relative  to  the  character  of  the  protein  material  or 
products  capable  of  absorption  have  been  greatly- 
influenced  naturally  by  the  ideas  concerning  the  nature 
and  extent  of  protein  digestion  prevalent  at  a  partic- 
ular period.  It  is  obvious  that  in  the  days  of  Liebig 
and  his  contemporaries  when  digestion  was  assumed 
to  be  little  more  than  a  process  whereby  proteins  were 
rendered  soluble  that  the  conception  of  the  absorption 
of  unchanged  protein  should  hold  sway.  Later,  after 
Kiihne  had  added  his  contributions  to  the  knowledge 
of  digestion,  theories  of  absorption  were  correspond- 
ingly modified.  Since  the  extent  of  formation  and  sig- 
nificance of  the  amino  acids  have  become  better  appre- 
ciated our  present-day  views  as  to  absorption  are  like- 
wise undergoing  modification. 

Absorption  from  the  Stomach 

A  great  deal  of  discussion  has  taken  place  regarding 
the  question  of  gastric  absorption  of  protein.  It  has 
been  asserted  by  Tobler  that  as  much  as  22  to  30  per 
cent  of  the  material  in  the  stomach  after  a  protein 


THE  ABSORPTION  OF  PROTEINS       47 

meal  is  absorbed.  London  and  his  co-workers,  on 
the  other  hand,  deny  that  any  absorption  takes  place. 
Abderhalden  with  Prym  and  London  have  apparently 
decided  the  question  in  the  negative  for  they  have 
demonstrated  that  amino  acids  fed  to  a  dog  with 
several  fistulae  almost  completely  pass  the  pylorus,  the 
first  absorption  occurring  in  the  duodenum.  Under 
normal  conditions,  therefore,  gastric  absorption  so 
far  as  protein  is  concerned  may  be  regarded  perhaps 
as  a  negligible  factor. 

Looked  at  from  another  viewpoint,  that  of  the 
present  with  its  modified  ideas  of  the  purpose  of  diges- 
tion, one  would  naturally  expect  little  or  no  absorption 
from  the  stomach.  If  the  view  is  correct  that  the  pur- 
pose of  alimentary  treatment  of  protein  is  to  hydrolyze 
this  substance  to  either  a  polypeptide  or  amino  acid 
stage  then  it  is  reasonable  to  assume  that  these  are  the 
products  absorbed,  rather  than  the  proteoses  or  pep- 
tones. Inasmuch  as  gastric  digestion  fails  to  decom- 
pose protein  to  the  stage  of  products  naturally  absorbed 
it  is  reasonable  to  assume  that  the  stomach  is  not  an 
organ  adapted  for  extensive  absorption  under  ordinary 
circumstances.  On  the  other  hand,  Folin  and  Lyman 
have  shown  conclusively  that  amino  acids,  Witte  pep- 
tone and  urea,  may  be  absorbed  from  the  stomach  when 
a  ligature  is  tied  around  the  pyloric  opening.  In  view 
of  these  conflicting  facts  one  is  hardly  justified  in 
making  a  positive  statement  as  to  the  importance  of 
the  stomach  in  the  absorption  of  nitrogenous  decom- 
position products. 


48  THE  AMINO  ACIDS 

Intestinal  Absorption 

From  extensive  experimentation  it  would  appear  that 
the  small  intestine  is  capable  of  absorbing  unchanged 
native  proteins  and  their  decomposition  products  the 
proteoses  peptones  and  amino  acids. 

Absorption  of  Undigested  Protein 

It  was  pointed  out  by  Voit  and  Bauer  in  1869  that 
undigested  proteins  such  as  serum  or  natural  egg  albu- 
min may  be  absorbed  by  the  small  intestine,  an  obser- 
vation which  has  been  repeatedly  confirmed  by  others. 
It  has  been  suggested  that  this  is  not  the  manner  in 
which  most  of  the  absorption  takes  place,  only  enough 
protein  being  absorbed  to  replace  worn-out  tissue,  the 
excess  being  oxidized  without  ever  having  entered  into 
the  tissue  metabolism  proper.  Various  explanations 
have  been  offered  to  account  for  the  fact  of  absorption 
of  undigested  protein.  The  most  obvious  assumption 
to  make  is  that  enzymes  must  have  been  present  in  the 
intestine  resulting  in  hydrolysis  to  amino  acids  and 
hence  their  absorption.  This  point,  however,  is  not 
valid  since  the  absorption  was  too  rapid  to  admit  the 
possibility  of  such  an  hypothesis.  Again,  it  has  been 
assumed  that  the  experimental  conditions  rendered  the 
intestinal  wall  unusually  permeable,  thus  allowing 
protein  to  pass.  It  is  possible  that  these  results  may 
later  be  explained  on  grounds  other  than  that  of  ab- 
sorption, for  Abderhalden,  Funk,  and  London  after 
introducing   excessive   amounts   of   protein   into   the 


THE  ABSORPTION  OF  PROTEINS       49 

intestine  failed  to  obtain  any  reaction  with  the  pre- 
cipitin test.  If  protein  were  actually  absorbed  un- 
changed in  its  natural  form  it  is  almost  incredible  that 
the  precipitin  test  failed  to  demonstrate  its  presence 
when  the  extreme  delicacy  of  this  reaction  is  recalled. 

What  Happens  to  Protein  Parenterally  Introduced? 

If  it  is  possible  for  unaltered  native  protein  to  be 
absorbed  by  the  intestinal  epithelium  is  it  capable  of 
supplying  the  nitrogenous  needs  of  the  body?  Or 
what  changes  does  it  undergo  after  absorption?  In 
attempts  to  answer  these  questions  endeavors  have 
been  made  to  follow  the  fate  of  native  proteins  intro- 
duced into  the  organism  with  avoidance  of  the  gastro- 
enteric tract.  For  many  years  it  has  been  accepted 
that  protein  introduced  parenterally  may  be  utilized  in 
part  at  least.  This  view  was  initiated  from  the  investi- 
gations of  Zuntz  and  v.  Mering  and  Neumeister.  Since 
then  it  has  been  repeatedly  confirmed  by  a  long  list 
of  observers.  Although  it  is  generally  admitted  that 
parenterally  introduced  protein  reappears  in  the  urine 
only  in  small  measure,  there  is  not  a  unanimity  of 
opinion  as  to  its  fate.  Even  though  the  intravenous 
injection  of  ^gg  albumin  fails  to  lead  to  a  large  output 
of  protein  in  the  urine  it  has  been  agreed  that  its 
failure  to  be  eliminated  by  the  kidney  is  no  evidence 
of  its  utilization  in  the  tissues.  In  such  an  argument 
one  might  assume  with  reason  that  the  protein  could 
be  excreted  through  the  bile,  be  poured  into  the  intes- 


50  THE  AMINO  ACIDS 

tine,  undergo  intestinal  digestion  and  eventually  be 
absorbed  in  the  form  of  protein  decomposition  pro- 
ducts. Experiments  to  test  this  point  have  been  carried 
out.  It  has  been  shown  that  when  a  solution  of  casein 
is  introduced  directly  into  the  blood  stream  a  small  part 
may  reappear  in  the  bile 

On  the  other  hand,  when  egg  white  or  serum  are 
injected  subcutaneously  into  dogs  and  goats  a  goodly 
portion  of  the  protein  may  be  eliminated  in  the  form 
of  non-coagulable  protein.  This  observation  would 
tend  to  demonstrate  the  non-utilization  of  the  injected 
protein  as  such  and  points  out  that  it  undergoes  a 
change  in  its  transit  through  the  organism.  In  the 
blood  also  a  non-coagulable  protein,  perhaps  a  proteose, 
is  detectable,  and  a  marked  increase  in  nitrogen  of 
the  urine — in  the  form  of  urea — is  apparent.  Indeed, 
nitrogen  equilibrium  may  be  maintained  under  these 
circumstances  when  animals  are  given  a  sufficiency  of 
carbohydrates. 

In  most  of  the  work  on  parenteral  absorption  of 
protein  the  material  introduced  did  not  possess  enough 
differentiation  from  other  body  proteins  to  distinguish 
it  from  them.  Borchardt  conceived  the  idea  of  inject- 
ing a  protein  with  peculiar  properties  and  chose  hemi- 
elastin ;  after  intravenous  injection  this  substance  was 
present  in  the  wall  of  the  small  intestine,  and  Borchardt 
concluded  that  the  protein  was  either  on  its  way  for 
excretion  by  the  gut  or  further  changes  by  the  intes- 
tinal juices  to  prepare  it  for  utilization  by  the  tissues, 
or  finally  had  found  its  way  into  the  intestine  by  way 


THE  ABSORPTION  OF  PROTEINS        51 

of  the  bile.  The  last  hypothesis  did  not  appear  likely 
however,  since  none  of  the  introduced  protein  could 
be  found  in  the  liver. 

From  the  foregoing  arguments  it  is  clear  that  the 
apparent  utilization  of  parenterally  introduced  protein 
may  be  disposed  of  in  at  least  three  ways :  1.  The 
direct  utilization  by  the  tissues,  for  which  there  is  little 
or  no  evidence.  2.  The  excretion  of  the  injected  mate- 
rial into  the  intestine  where  it  is  subjected  to  the 
action  of  digestive  enzymes,  finding  its  way  either 
directly  into  the  intestine  or  indirectly  through  the 
bile.  There  is  little  doubt  that  a  certain  portion  of 
the  injected  material  is  treated  in  this  manner.  3.  The 
transformation  of  the  native  protein  in  the  tissues 
into  smaller  fractions  such  as  proteoses,  peptones  or 
amino  acids.  Heilner  has  suggested  that  the  utilization 
of  parenterally  introduced  protein  is  induced  by  the 
generation  of  a  specific  enzyme  capable  of  bringing 
about  a  hydrolysis. 

In  the  last  suggestion  it  is  probable  that  we  have 
the  true  explanation  for  the  phenomenon  under  dis- 
cussion for  Abderhalden  and  his  co-workers  have 
demonstrated  that  the  parenteral  introduction  of  native 
protein  or  of  peptone  gives  to  the  blood  serum  of  the 
animal  the  power  of  decomposing  these  substances,  and 
this  power  is  destroyed  by  heating  to  60°  to  65°  C. 
Over-feeding  by  mouth  confers  upon  the  blood  serum 
the  same  property.  The  acquisition  of  this  power  on 
the  part  of  the  blood  serum  may  be  regarded,  as 
Heilner  suggested,  as  a  generation  of  an  enzyme  or 


52  THE  AMINO  ACIDS 

it  may  be  possible  that  the  transport  of  the  foreign 
material  through  the  tissue  has  carried  with  it  into  the 
blood  enzymes  already  existing  in  other  parts  of  the 
organism.  Be  this  as  it  may,  it  is  very  probable  that 
protein  retained  in  the  body  after  parenteral  intro- 
duction or  even  perhaps  after  absorption  from  the 
intestinal  canal  without  profound  disintegration  even- 
tually undergoes  decomposition  into  simpler  products 
after  reaching  the  blood  stream.  It  would  appear  from 
this  statement,  therefore,  that  the  tissues  must  prefer, 
to  say  the  least,  their  pabulum  in  the  form  of  relatively 
simple  compounds  rather  than  as  complex  molecules 
like  the  native  proteins. 

Absorption  of  Proteoses  and  Peptones 

It  was  early  discovered  that  peptone  left  in  contact 
with  the  living  intestinal  wall  disappeared  or  at  least 
failed  to  show  its  characteristic  reactions.  From  these 
observations  Hofmeister  formulated  the  theory  that 
the  peptones  were  taken  up  by  the  leucocytes  of  the 
intestine  after  absorption  and  by  them  transformed 
into  protein  and  distributed  to  the  tissues.  This 
hypothesis  failed  to  meet  with  the  approval  of  Heiden- 
hain,  who,  although  believing  in  the  conversion  of  pep- 
tone to  protein,  assigned  to  the  intestine  itself  the 
important  role  of  this  transformation.  In  confirmation 
of  the  correctness  of  this  idea  may  be  cited  the  experi- 
ments of  Hofmeister  and  Neumeister  who  demon- 
strated that  peptone  introduced  directly  into  the  blood 


THE  ABSORPTION  OF  PROTEINS       53 

was  treated  as  so  much  waste  material  being  eliminated 
by  the  kidneys.  Moreover,  he  also  failed  to  find  any 
trace  of  peptone  in  the  blood  or  lymph  of  animals  at 
the  height  of  digestion. 

The  experiments  cited  above  failed  to  throw  any 
light  upon  the  fate  of  the  peptone,  beyond  its  dis- 
appearance. In  later  experiments  Neumeister  showed 
the  presence  of  leucine  and  tyrosine  in  the  intestine 
after  introduction  of  peptone,  thus  indicating  a  further 
decomposition  of  the  peptone.  Even  though  it  might 
be  accepted  that  peptone  placed  in  the  intestine  under- 
goes a  further  breakdown  to  amino  acids  there  still 
existed  no  proof  that  the  amino  acids  were  absorbed 
as  such.  It  is  possible  to  assume  a  synthesis  of  the 
amino  acids  back  into  protein  in  the  act  of  absorption 
through  the  intestinal  wall.  An  example  of  such  a 
reaction  is  found  in  the  digestion  of  fat  where  fat  is 
split  into  fatty  acids  or  soap  and  glycerol  and  regener- 
ated as  fat  during  the  process  of  absorption. 

It  was  Cohnheim's  attempt  to  isolate  this  hypotheti- 
cal protein  from  the  intestinal  wall  which  led  to  his 
discovery  of  the  enzyme  erepsin  already  considered  the 
action  of  which  has  had  a  tendency  toward  filling  in 
some  of  the  gaps  in  our  conception  of  the  nature  of 
intestinal  digestion  and  absorption.  Although  it  had 
been  recognized  for  many  years  that  amino  acids  are 
formed  in  intestinal  digestion  they  were  regarded  as 
by-products  and  quite  unimportant.  As  a  result  of 
the  discovery  of  erepsin  the  purpose  of  the  formation 
of  amino  acids  first  received  its  due  recognition. 


54  THE  AMINO  ACIDS 

That  all  of  the  older  investigators  did  not  regard  the 
direct  absorption  of  protein  or  even  of  such  large 
molecules  as  peptone  as  essential  for  nutrition  may  be 
seen  from  the  view  formulated  by  Salkowski  and 
Leube.  According  to  this  suggestion  leucine  may  be 
regarded  as  a  substance  capable  after  absorption  of 
being  built  up  into  protein  and  therefore  leucine  might 
be  looked  upon  as  a  stage  in  protein  regeneration. 
Against  such  a  view,  however,  stood  the  fact  that  the 
amino  acids  were  not  at  that  time  demonstrable  in  the 
blood  or  lymph. 

From  the  numerous  researches  carried  through  con- 
cerning the  absorption  of  proteoses  and  peptones  from 
the  intestines,  the  conclusion  may  be  drawn  that 
although  these  proteins  disappear  when  placed  in  con- 
tact with  the  intestinal  mucosa,  there  is  no  evidence  of 
their  absorption  as  such  for  they  can  be  found  neither 
in  the  blood  nor  in  the  lymph.  On  the  other  hand, 
inasmuch  as  their  disappearance  from  the  intestine  is 
coincident  with  the  appearance  of  amino  acids,  an 
enzyme  being  furnished  which  specifically  effects  such 
a  transformation,  it  is  probable  that  these  proteins  are 
absorbed  only  in  the  form  of  amino  acids. 

The  Absorption  of  Amino  Acids 

As  has  been  stated  previously  the  presence  of  amino 
acids  in  the  small  intestine  has  long  been  known.  Their 
absorption  therefrom,  however,  has  been  a  matter  of 
conjecture.     Inasmuch  as  their  presence  in  the  blood 


THE  ABSORPTION  OF  PROTEINS       55 

or  lymph  could  not  be  detected  the  theory  of  their 
synthesis  to  protein  coincident  with  their  absorption 
was  promulgated.  To  Folin  we  owe  the  first  indirect 
proof  of  the  absorption  of  amino  acids  from  the  intes- 
tines. By  a  set  of  delicate  methods  adapted  for  the 
partition  of  different  forms  of  nitrogen  he  has  suc- 
ceeded in  demonstrating  an  appreciable  increase  in  the 
"non-protein"  portion  of  the  blood,  after  introduction 
of  amino  acids  into  a  loop  of  intestine.  Although  the 
amino  acids  themselves  were  not  isolated  the  non- 
protein fraction  of  the  blood  was  so  significant  as  to 
leave  no  doubt  of  amino  acid  absorption.  Van  Slyke 
and  Meyer  strongly  fortified  the  view  by  the  use  of  a 
different  method.  The  quantity  of  amino  acids  in  the 
circulation  at  one  time  is  so  small  as  to  have  escaped 
detection  by  methods  previously  in  use.  It  remained 
for  Abderhalden  to  demonstrate  the  presence  of  amino 
acids  in  the  blood  under  normal  circumstances  and 
actually  to  isolate  them.  This  he  accomplished  by 
employing  fifty  liter  lots  of  blood.  From  such  large 
volumes  he  succeeded  in  separating  and  identifying 
proline,  leucine,  valine,  aspartic  acid,  glutamic  acid, 
alanine,  glycocoU,  arginine,  histidine,  and  lysine.  In 
no  instance  did  he  obtain  more  than  0.5  gram  of  any 
one  substance.  The  absorption  of  amino  acids  as  such 
is,  therefore,  an  assured  fact. 

Absorption  from  the  Large  Intestine 

Recalling  to  mind  the  heterogenous  mixture  of  sub- 
stances that  may  reach  the  large  intestine  one  at  once 


66  THE  AMINO  ACIDS 

realizes  the  great  number  of  compounds  that  may  find 
their  way  into  the  blood  stream.  Leaving  out  of  con- 
sideration any  residue  of  undigested  protein  we  may 
confine  our  attention  to  the  possibilities  of  proteose  and 
peptones,  the  amino  acids  and  the  derivatives  of  the 
latter.  The  evidence  of  the  absorption  of  proteose  and 
peptone  derivatives  is  decisive  since  Folin  and  Denis 
have  demonstrated  an  increase  in  the  non-protein 
nitrogen  of  the  blood  after  placing  Witte  peptone  in  a 
ligatured  loop  of  the  large  intestine.  The  absorption 
from  the  large  intestine,  however,  is  much  slower  than 
obtains  in  the  small  intestine.  In  a  similar  manner 
Folin  and  Denis  have  observed  the  absorption  of  dif- 
ferent amino  acids  and  urea.  Throughout  the  entire 
intestinal  canal,  therefore,  the  absorption  of  amino 
acids  may  be  regarded  as  a  normal  process. 

The  absorption  of  the  well-known  typical  products 
of  putrefaction  needs  only  brief  description  since  their 
fate  has  long  been  recognized.  Absorption  of  indole, 
skatole,  phenol,  cresol,  etc.,  is  certain  since  their 
addition  products  are  found  in  the  urine.  Thus  indole 
is  absorbed,  carried  to  the  liver  through  the  portal 
vein,  oxidized  to  hydroxy  indole  (indoxyl),  combined 
with  sulphuric  acid  and  eventually  is  eliminated  as  the 
potassium  salt,  indican — its  amount  being  indicative 
of  the  extent  of  intestinal  putrefaction.  Or  indole  may 
be  combined  in  part  after  oxidation  with  glycuronic 
acid  and  be  excreted  as  a  glycuronate.  Phenol  and 
cresol  may  likewise  be  eliminated  in  the  urine  as 
ethereal  sulphates. 


THE  ABSORPTION  OF  PROTEINS        57 

The  fate  of  the  amines  formed  in  putrefaction  is  also 
fairly  well  established,  at  least  in  certain  instances. 
Thus,  for  example,  it  is  known  that  the  amine  formed 
from  tyrosine,  p .  oxyphenylethylamine  in  passing  the 
organism  is  transformed  to  and  excreted  as  p.oxy- 
phenylacetic  acid. 

References  to  Literature 

Ahderhalden:  Zeitschrlft  fiir  physiologische  Chemie.  1913, 
88 y  p.  478.    [Amino  acids  in  blood.] 

Cathcart:  The  Physiology  of  Protein  Metabolism.    1912. 

Folin  and  Denis:  Journal  of  Biological  Chemistry.  1912,  ii, 
p.  87  and  p.  161;  1912,  I2,  p.  141  and  p.  253.  [Fate  of 
digestion  products.] 

Hammarsten:  Text  Book  of  Physiological  Chemistry.    1914. 

Van  Slyke  and  Meyer:  Journal  of  Biological  Chemistry. 
1912,  12,  p.  399;  1913-1914,  i6,  p.  187,  p.  197,  p.  213,  and 
p.  231.    [Fate  of  digestion  products.] 


CHAPTER  IV 

IN  WHAT  FORM  DOES  INGESTED  PROTEIN 
ENTER  THE  CIRCULATION? 

Our  conception  of  the  nature  of  metabolic  processes 
in  the  tissues  will  be  more  or  less  modified  by  the  view 
accepted  concerning  the  degree  and  character  of  dis- 
integration of  protein  induced  in  the  alimentary  tract 
and  the  form  of  the  products  absorbed.  This  being  so 
a  consideration  of  the  hypotheses  that  have  been  ad- 
vanced relative  to  the  fate  of  protein  after  its  dis- 
appearance from  the  gastro-enteric  tract  is  now  in 
order.  This  fate  of  ingested  protein  has  been  ex- 
plained in  at  least  four  different  ways,  namely : 

1.  The  proteins  are  absorbed  with  little  or  no 
chemical  change  and  are  taken  up  by  the  tissues  and 
incorporated  into  them.  In  a  previous  chapter  it  has 
been  noted  that  native  protein  may  be  absorbed  as 
such  at  times  and  fail  to  reappear  leading  to  the  infer- 
ence of  utilization  by  the  tissues.  This,  however, 
cannot  be  accepted  as  the  usual  procedure  for  all  the 
food  protein.  Once  in  the  blood  stream  as  shown  by 
parenteral  introduction  protein  utilization  apparently 
occurs  through  the  intervention  of  enzymes  which  sud- 
denly make  their  appearance  in  the  circulation. 


PROTEIN  AND  CIRCULATION  59 

2.  The  proteins  of  the  food  are  hydrolyzed  and  the 
products  are  absorbed  and  carried  to  the  tissues. 

3.  The  digestion  products  are  synthesized  into 
serum  protein  by  the  intestinal  wall  during  the  act  of 
absorption,  and  the  serum  protein  serves  as  pabulum 
for  the  tissues. 

4:.  Deamination  of  the  digestion  products  occurs 
previous  to  their  entrance  into  the  circulation.  Since 
it  is  impossible  to  accept  the  hypothesis  that  unchanged 
protein  is  the  form  in  which  ingested  protein  is  usually 
absorbed  the  next  natural  inference  is  that  the  pro- 
teoses and  peptones  are  absorbed  directly  into  the  blood 
and  conveyed  to  the  tissues.  In  attempting  to  deter-^* 
mine  the  correctness  of  this  hypothesis  the  query  has 
arisen 

Are  Proteoses  and  Peptones  Present  in  the 

Blood  ? 

In  spite  of  the  discovery  of  erepsin  by  Cohnheim 
and  the  consequent  improbability  of  proteoses  and 
peptone  representing  the  usual  form  of  final  digestion 
products,  some  modern  investigators  have  clung  to  the 
idea  that  it  is  in  the  form  of  proteoses  and  peptones 
that  protein  is  absorbed.  This  view  is  based  upon  a 
number  of  investigations  from  which  the  assertion  has 
been  made  that  proteoses  and  peptones  are  present  in 
the  blood  stream.  The  work  of  Neumeister  led  to  the 
conclusion  that  proteoses  and  peptones  are  not  found 
in  the  blood  and  it  was  not  until  1903  through  the 


60  THE  AMINO  ACIDS 

observations  of  Embden  and  Knoop  that  any  doubt 
of  Neumeister's  view  was  entertained.  In  experiments 
designed  to  show  the  fate  of  proteoses  and  peptones 
when  brought  into  contact  with  the  intestinal  wall 
Embden  and  Knoop  were  able  to  show  the  presence  in 
the  blood  of  substances  having  the  properties  of  pro- 
teoses and  peptones.  They  held  that  the  mucous  mem- 
brane of  the  intestine  neither  synthesized  these  sub- 
stances to  a  larger  molecule,  as  for  example  to  a  coagu- 
lable  protein,  nor  were  they  hydrolyzed  to  the  amino 
acid  stage  but,  on  the  contrary,  absorption  took  place 
directly.  The  results  of  Embden  and  Knoop  were  con- 
firmed by  some  observers  and  discredited  by  others. 
Schumm  for  example  was  always  unable  to  find  a  trace 
of  proteose  in  the  blood  both  under  normal  and  ab- 
normal conditions  of  health.  Abderhalden  and  his  co- 
workers maintained  that  the  substances  giving 
reactions  for  proteoses  and  peptones  were  present 
because  of  imperfect  methods  employed  in  the  separa- 
tion of  the  coagulable  protein  from  the  blood. 

It  has  been  pointed  out  by  others,  however,  that  there 
is  a  possibility  of  the  presence  in  the  blood  of  a  protein 
naturally  non-coagulable  which  would  still  give  the 
reaction — the  biuret  test — significant  under  the  experi- 
mental conditions  for  the  presence  of  proteose  or  pep- 
tone. Zanetti  described  such  a  protein  in  the  blood 
and  found  that  it  belonged  to  the  group  of  mucoids. 
Among  others,  Howell  believes  in  the  existence  in  the 
blood  of  a  protein  possessing  some  of  the  character- 
istics of  the  proteoses  and  peptones,  for  example,  non- 


PROTEIN  AND  CIRCULATION  61 

coagulability,  which  is  not,  however,  identical  with 
these  substances.  Bywaters  has  concluded  that  this 
body  is  a  substance  called  by  him  sero-mucoid.  In  spite 
of  these  views  Bergmann  and  Langstein  and  Kraus 
assert  that  small  amounts  of  true  proteose  are  present 
constantly.  After  feeding  elastin  Borchardt  claimed  to 
find  elastin  proteose  in  the  blood  stream.  Upon  repe- 
tition of  this  work  of  Borchardt,  Abderhalden  and 
Ruehl  failed  to  give  it  confirmation.  The  explanation 
of  Abderhalden  and  Oppenheimer  that  imperfect  sepa- 
ration of  coagulable  protein  is  responsible  for  the 
proteose  test  was  denied  by  Freund  who  maintained 
that  the  method  employed  by  Abderhalden  not  only 
precipitated  the  coagulable  protein  but  the  proteose 
also. 

From  the  foregoing  brief  review  of  only  a  few  of  the 
investigations  carried  through  for  the  decision  of  the 
problem  it  is  apparent  that  the  whole  question  is  in  a 
chaotic  state  of  contradictions  and  that  a  positive 
answer  cannot  be  given.  It  may  at  least  be  said  that 
positive  proof  of  the  presence  of  proteoses  and  pep- 
tones is  still  lacking.  Perhaps  one  of  the  most  con- 
vincing arguments  against  the  existence  in  the  blood  of 
proteoses  and  peptones  is  derived  from  the  work  of 
Abderhalden  and  Pincussohn.  They  have  demon- 
strated that  just  as  with  the  parenteral  introduction 
into  the  body  of  native  protein  so  with  proteose  injec- 
tion there  is  a  development  in  the  blood  plasma  of  an 
enzyme  capable  of  causing  its  disintegration  to  smaller 
molecules.     Such  an  enzyme  is  not  present  in  the 


62  THE  AMINO  ACIDS 

blood  under  ordinary  conditions.  If  proteoses  were 
present  normally  in  the  blood  it  is  probable  that  this 
enzyme  would  also  be  a  normal  constituent  of  the 
blood. 

The  Synthesis,  or  Regeneration  of  Protein,  by 
THE  Intestine 

If  it  is  accepted  that  protein  is  not  absorbed  in  the 
form  of  proteoses  or  peptones  the  query  naturally 
arises,  In  what  form  is  it  absorbed  ?  An  answer  to  this 
question  must  necessarily  determine  also  the  place  of 
protein  regeneration  so  long  as  the  conception  of  nutri- 
tion obtains  that  metabolic  changes  in  the  organism 
demand  the  formation  of  new  material  to  replace  that 
broken  down.  If  one  maintains  that  protein  gets  into 
the  blood  stream  in  the  form  of  a  molecule  larger  than 
the  amino  acid,  proteose  or  peptone  molecule,  it  is  self- 
evident  that  the  intestine  must  be  regarded  as  capable 
of  synthesizing  amino  acids  to  protein.  On  the  other 
hand  if  amino  acids  are  regularly  present  in  the  sys- 
temic circulation,  the  place  of  protein  regeneration 
must  be  relegated  to  the  cellular  elements  of  the  dif- 
ferent tissues. 

Ahderhalden.  In  the  past  various  theories  have  been 
maintained.  In  view  of  the  failure  to  find  amino  acids 
in  the  blood,  Abderhalden  put  forward  the  view  that 
the  intestinal  wall  possessed  the  power  during  the  act 
of  absorption  to  synthesize  the  amino  acids  to  proteins, 
probably  serum  proteins,  to  meet  the  needs  of  the 


PROTEIN  AND  CIRCULATION  63 

organism's  requirements.  The  necessity  for  prelimi- 
nary extensive  disintegration  of  the  food  protein  has 
been  offered  as  follows :  "Every  species  of  animal — in 
fact  every  individual — has  its  specifically  constituted 
tissues  and  cells.  If  the  diet  was  always  the  same,  the 
formation  of  the  tissues  might  bear  some  close  relation 
to  the  components  of  the  food.  The  diet  varies,  how- 
ever, and,  especially  in  the  case  of  human  beings  and 
the  omnivora,  is  exceedingly  diverse  in  nature  and  to 
make  its  organism  independent  of  the  outer  world  in 
the  matter  of  food  taken,  it  disintegrates  the  nutrient 
it  receives,  and  utilizes  those  components  which  may  be 
of  service  to  it  in  building  up  new  complexes." 

Objections  to  this  theory  have  been  summarized  by 
Cathcart  as  follows :  "The  view  that  the  tissue  proteins 
differ  from  one  another,  that  they  are  specific  bodies 
of  definite  constitution,  and  that,  therefore,  each  re- 
quires a  different  amount  and  supply  of  building  mate- 
rial is  gradually  being  accepted.  Abderhalden  himself 
accepts  this.  What  end  then  is  served  in  having  a 
single  uniform  pabulum  formed  when  the  demand  is 
so  varied?  This  is  all  the  more  questionable  when  it 
is  remembered  that  there  is  no  indubitable  evidence 
which  shows  that  one  amino  acid  can  be  converted 
into  another.  Further,  the  belief  is  gradually  gaining 
ground,  as  regards  the  protein  requirements  of  the 
organism,  that  it  is  not  so  much  the  actual  quantity  as 
the  quality  of  the  protein  supplied  in  the  food,  which 
is  of  importance.  If  the  material  supplied  be  uniform 
it   necessitates   a   fresh   breakdown   by   each   tissue, 


64  THE  AMINO  ACIDS 

perhaps  by  each  individual  cell.  Although  the  tissues 
all  probably  possess  this  power  of  breaking  down  pro- 
tein material  by  means  of  their  intracellular  proteolytic 
enzymes,  still  the  extra  work  involved  seems  to  nega- 
tive the  immediate  resynthesis  hypothesis,  especially 
when  the  hypothesis  of  the  circulating  digestion  pro- 
duct postulates  the  presence  of  the  individual  food 
material  in  the  blood.  As  already  remarked,  the  mere 
failure  to  detect  these  products  in  the  blood  does  not 
give  adequate  reason  for  concluding  that  they  are  not 
present.  The  tissues  certainly  do  not  break  dov/n  in 
regular  sequence,  nor  are  they  left  to  fall  to  pieces  for 
lack  of  repair  material.  Repair  is  among  the  most 
active  functions  of  all  tissues.  Must,  then,  a  tissue 
of  highly  complex  structure  keep  destroying  and  digest- 
ing plasma,  picking  out  from  the  debris  the  nuclei 
which  it  requires  and  letting  the  rest  go  ?  (Why,  and 
this  destruction  is  admitted  by  Abderhalden,  are  the 
superfluous  amino  acids  not  found  in  the  blood?) 
What  happens,  for  instance,  in  the  case  of  connective 
tissues  with  their  demand  for,  say,  glycine,  where  the 
food  supply  is  not  over-abundant  as  the  circulation  is 
poor,  and  the  tissues  not  very  suited  for  lymph  per- 
fusion? It  will  not  do  merely  to  say  that  there  is  no 
great  breakdown  of  material  here.  Pfliiger,  in  an  inter- 
esting paper  in  which  he  combated  this  immediate 
resynthesis  hypothesis,  ascribed  to  the  cells  of  the 
intestinal  wall,  with  regard  to  the  protein  synthesis, 
the  same  capacity  as  the  cells  of  all  tissues,  but  denied 
that  the  synthesis  of  protein  for  the  whole  organism 


PROTEIN  AND  CIRCULATION  65 

was  carried  out  there.  He  held  that  such  a  hypothesis 
was  contrary  to  all  existent  knowledge  of  physiological 
assimilation." 

One  may  query  to  what  extent  does  immediate 
resynthesis  take  place  ?  Are  all  the  digestion  products 
transformed  into  coagulable  protein  or  are  some  se- 
lected and  others  rejected  in  part?  The  questions 
cannot  be  answered  by  the  supporters  of  Abder- 
halden*s  theory.  On  the  other  hand,  the  intestine  evi- 
dently is  capable  of  exerting  a  marked  selective  action 
as  to  the  type  and  amount  of  amino  acid  it  shall  absorb. 
The  experiments  of  Abderhalden  and  his  co-workers 
have  indicated  this.  They  fed  gliadin  to  polyfistular 
animals  and  observed  as  the  material  traversed  the 
gastro-enteric  tract  that  tyrosine  disappeared  from  the 
intestine  whereas  glutamic  acid  steadily  increased  in 
amount. 

Freund,  The  idea  of  Freund  is  somewhat  similar 
to  the  hypothesis  advanced  by  Abderhalden  except 
that  he  ascribes  to  the  liver  an  important  role  in  the 
subsequent  breakdown  of  the  protein.  The  protein 
digestion  products  are  assumed  by  Freund  to  travel 
the  portal  circulation  in  the  form  of  pseudo-globulin. 
The  liver  is  unable  to  properly  decompose  protein 
unless  it  has  first  entered  the  blood  stream  by  way  of 
the  intestine.  This  hypothesis  carries  with  it  the  sug- 
gestion that  the  parenteral  utilization  of  protein  must 
be  carried  out  through  aid  from  the  intestine,  that  the 
protein  is  excreted  from  the  blood  into  the  intestine, 
undergoes  digestion,  the  products  are  absorbed,  and 


66  THE  AMINO  ACIDS 

during  the  act  of  absorption  are  polymerized,.  This 
suggestion  has  been  tested  in  different  ways.  One 
might  expect  that  protein  parenterally  introduced  into 
dogs,  the  intestine  of  which  had  been  removed,  would 
reappear  in  the  urine.  Korosy  has  failed  to  find  more 
than  traces  of  protein  after  parenteral  injections  of 
protein  into  animals  without  an  alimentary  tract.  These 
observations  tend  to  show  that  intestinal  preparation 
of  protein  cannot  be  regarded  as  essential.  The  prob- 
lem was  attacked  in  another  way  by  Abderhalden  and 
London.  They  attempted  to  determine  the  excretion  of 
protein  into  the  intestine  of  polyfistular  animals  after 
parenteral  introduction  but  failed  to  obtain  any  evi- 
dence of  such  a  reaction.  On  the  other  hand,  the  excre- 
tion of  substances  into  the  intestine  after  parenteral 
injection  is  known.  Thus,  Abderhalden  and  Slavu 
have  shown  that  iodine  may  find  its  way  into  the 
intestine  when  iodine-polypeptide  combinations  are 
injected  subcutaneously. 

Hofmeister.  It  was  the  view  of  Hofmeister  that  the 
leucocyte  is  intimately  associated  with  protein  re- 
generation. The  idea  undoubtedly  originated  from  the 
marked  leucocytosis  which  occurs  after  meals  and 
Hofmeister  thought  that  peptone  after  absorption  was 
changed  in  some  unknown  manner  into  protein  by  the 
leucocytes  or  else  through  the  agency  of  adenoid  tissue. 
Later,  the  lymphocyte  was  selected  as  the  specific  form 
of  leucocyte  responsible  for  protein  synthesis.  This 
theory,  however,  finds  few  supporters  today.  Perhaps 
the  best  criticism  of  the  leucocyte  synthesis  theory  has 


PROTEIN  AND  CIRCULATION  67 

been  offered  by  Halliburton.  "He  pointed  out  that  the 
number  of  the  lymphocytes  was  not  commensurate 
with  the  work  to  be  done.  He  calculated  that  a  man 
of  eighty  kilos  had  about  four  kilos  of  blood  of  which 
some  40  per  cent  was  in  the  form  of  corpuscles,  that  is 
about  1600  grams.  Now  as  the  ratio  of  white  cor- 
puscles is  about  1 :  500  it  means  that  about  3.2  grams  of 
leucocytes  are  present.  Of  this  amount  lymphocytes 
form  at  most  30  per  cent,  and  therefore  in  the  blood 
there  would  be  about  one  gram  of  lymphocytes.  If 
this  amount  w^ere  doubled  during  digestion  *it  is  diffi- 
cult to  see  how  two  grams  of  lymphocytes  can  tackle 
the  enormous  burden  which  every  meal  must  impose 
upon  them.' "     (Cathcart.) 

What  Is  the  Evidence  for  the  Synthesis  of 

Protein  ? 

The  Synthetic  Action  of  the  Gastric  and  Intestinal 
Mucous  Membranes. 

Hofmeister  has  ascribed  to  the  stomach  mucous 
membrane  the  property  of  synthesizing  protein  from 
proteoses.  An  outline  of  his  experiment  follows — 
at  the  height  of  digestion  a  dog  was  killed  and  its 
stomach  and  contents  divided  equally  into  two  parts. 
One  part  was  immediately  placed  in  boiling  water  to 
stop  all  enzyme  and  cellular  activity  and  the  other 
portion  was  placed  in  an  incubator  for  a  period  of 
two  hours.  The  amount  of  proteose  and  peptone 
present  in  each  part  was  then  determined.    In  the  por- 


68  THE  AMINO  ACIDS 

tion  placed  in  the  incubator,  there  was  an  almost  com- 
plete disappearance  of  proteose  and  peptone,  which  of 
course  could  not  be  ascribed  to  further  decomposition 
since  gastric  juice  does  not  hydrolyze  proteoses  and 
peptones  to  amino  acids,  at  least  during  such  a  short 
period.  The  conclusion  drawn  was  that  the  proteoses 
and  peptones  disappeared  because  of  their  synthesis 
to  protein.  In  other  experiments  Hofmeister  demon- 
strated that  the  intestine  possesses  the  same  property. 
Glaessner  repeated  and  confirmed  Hofmeister's  inves- 
tigation. On  the  other  hand,  Embden  and  Knoop 
failed  to  find  any  evidence  of  protein  synthesis.  They 
employed  the  normal  intestine  and  also  the  intestine 
from  which  pancreatic  juice  was  excluded  by  ligature 
of  the  duct.  The  evidence  for  resynthesis  of  protein 
in  the  gastric  or  intestinal  mucous  membranes  is  not 
convincing  and  one  must  obtain  other  than  negative 
evidence  before  the  idea  of  such  a  protein  resynthesis 
can  be  accepted. 

Plasfein  Formation.  It  has  been  observed  repeatedly 
that  when  solutions  of  proteoses  are  brought  into  con- 
tact with  rennin  a  precipitate  called  plastein  forms. 
Various  views  as  to  its  formation  have  been  held.  It 
has  been  assumed  by  some  that  plastein  is  a  new  syn- 
thetic product  formed  from  the  proteoses — a  new  pro- 
tein, by  others  a  resynthesis  of  the  proteoses  to  the 
original  protein  from  which  they  were  derived,  and  by 
still  others  as  a  digestion  product  on  its  way  to  com- 
plete solution.  The  results  of  the  most  searching 
investigations  concerning  the  nature  of  plastein  incline 


PROTEIN  AND  CIRCULATION  69 

one  to  the  belief  that  this  substance  is  of  the  nature 
of  proteose  rather  than  that  of  a  complete  protein  so 
that  plastein  formation  affords  little  or  no  evidence  for 
the  support  of  the  existence  of  protein  synthesis. 

The  Synthetic  Action  of  Pepsin  and  Trypsin 

It  has  been  demonstrated  by  A.  E.  Taylor  that  by 
the  long  continued  action  of  trypsin  of  the  clam  liver 
upon  concentrated  protamine  digestion  products  a 
reformation  of  protamine  takes  place.  The  quantity 
reformed  is  very  small  in  comparison  with  the  original 
amount  of  protamine  digestion  products.  In  a  similar 
manner  Robertson  has  found  the  synthesis  of  a  para- 
nuclein  by  the  action  of  pepsin  upon  concentrated 
casein  digestion  products.  These  results  lead  to  the 
suggestion  that  trypsin  and  pepsin  may  possess  a  two- 
fold action,  a  disintegrative  influence  and  a  synthetic 
action  in  accord  with  the  idea  of  the  reversibility  of 
enzymes.  If  the  synthetic  action  in  the  intestine  is  as 
slow  as  that  shown  in  the  experiments  just  cited  little 
value  can  be  assigned  to  them  as  aids  in  the  regenera- 
tion of  protein  in  the  body  for  the  influence  could  be 
observed  only  after  the  influence  had  continued  for 
several  months. 

The  evidence  for  the  synthesis  of  protein  in  the 
intestinal  wall  is  all  of  an  indirect  nature.  If  the 
adherents  of  that  theory  could  demonstrate  an  increase 
of  protein  in  the  blood  after  an  ingestion  of  protein 


70  THE  AMINO  ACIDS 

their  argument  might  be  established.    This  they  have 
failed  to  do. 

Deamination 

The  failure  to  demonstrate  the  presence  of  amino 
acids  in  the  blood  of  the  higher  animals  during  diges- 
tion led  to  the  conception  that  the  amino  acids  are 
deaminated,  that  is,  ammonia  is  split  off  while  passing 
the  intestinal  wall,  this  deamination  being  regarded  as 
the  first  stage  in  the  catabolism  of  the  amino  acids. 
This  possibility  was  suggested  by  the  work  of  Cohn- 
heim  upon  certain  of  the  lower  forms  of  animal  life 
in  which  he  showed  the  giving  off  of  ammonia  by  the 
intestine  after  addition  of  amino  acids,  and  derived 
support  from  the  older  work  of  Nencki  and  others 
who  showed  that  the  ammonia  content  of  the  portal 
blood  was  greater  than  that  of  the  arterial  during 
digestion.  It  has  been  assumed  that  as  result  of  this 
process  of  deamination  the  ammonia  split  off  is  trans- 
formed by  the  liver  into  urea  and  so  quickly  eliminated 
by  the  kidneys.  Such  a  view  has  been  adopted  as 
explanatory  for  the  long  known  rapid  rise  in  urea 
excretion  following  protein  ingestion. 

It  has  been  shown  by  Lang  that  deamination  is  a 
property  of  a  great  many  tissues  of  the  body  but  it  is 
probable  that  certain  of  them  possess  a  selective  action 
in  this  respect  for  some  tissues  deaminate  certain  of  the 
amino  acids  much  more  readily  than  others.  In  par- 
ticular the  intestine  and  liver  seem  to  possess  this 
action  in  a  high  degree.    To  the  liver  a  great  import- 


PROTEIN  AND  CIRCULATION  71 

ance  has  been  attached  as  a  deaminating  agent  and 
during  recent  years  discussion  of  the  so-called  defect- 
ive or  insufficient  deamination  in  a  series  of  pathologi- 
cal conditions  has  come  into  vogue. 

In  accordance  with  this  idea  amino  acids  have  been 
administered  as  a  test  for  the  functional  activity  of 
the  liver.  Glaessner  has  shown  that  normal  liver  tissue 
is  capable  of  transforming  definite  amounts  of  specific 
amino  acids  into  urea.  In  a  series  of  experiments  he 
has  shown  that  in  various  diseased  conditions  of  the 
liver,  such  as  fatty  liver,  in  syphilis,  cirrhotic  liver,  and 
a  phosphorus  poisoned  liver  a  failure  to  convert  amino 
acids  into  urea  and  a  consequent  output  of  amino  acids 
in  the  urine  took  place. 

That  deamination  is  undoubtedly  an  important  intra- 
cellular activity  may  be  derived  from  a  series  of  experi- 
ments in  which  amino  acids  have  been  fed  and  their 
fate  determined.  Thus  with  arginine  most  of  the 
nitrogen  reappears  as  urea.  Probably  through  the 
intervention  of  the  enzyme,  arginase,  a  splitting  of 
arginine  into  urea  and  orinthine  occurs  and  by  deami- 
nation of  the  latter  more  urea  is  formed.  Again, 
after  administration  of  alanine,  lactic  acid  in  the  urine 
has  been  observed.  With  the  purines  also  it  may  be 
shown  that  a  splitting  off  of  ammonia  occurs.  One 
may  accept  without  hesitation  that  the  function  of 
deamination  is  an  important  activity  of  cell  life.  The 
contention  that  certain  organs  or  tissues  possess  this 
function  more  specifically  than  others  has  been  a  matter 
of  controversy.    The  role  played  by  the  intestine  in 


72  THE  AMINO  ACIDS 

this  regard  is  especially  to  be  considered,  correlated 
as  it  has  been  with  the  explanation  of  the  form  in 
which  protein  digestion  products  are  absorbed. 

The  recent  observations  of  Folin  and  Denis  and 
others  have  rendered  untenable  the  hypothesis  that 
deamination  by  the  intestine  is  the  first  stage  in  the 
catabolism  of  amino  acids.  They  demonstrated  that 
during  the  absorption  of  amino  acids  from  the  intes- 
tine there  was  no  increase  in  ammonia  or  urea  of  the 
blood  and  they  further  showed  that  the  ammonia  of 
the  portal  blood  is  produced  in  large  measure  by  the 
products  of  putrefaction  in  the  large  intestine.  The 
retirement  of  the  theory  of  intestinal  deaminization 
to  account  for  the  apparent  absence  of  amino  acids  in 
the  blood  carries  with  it  also  the  untenability  of  the 
idea  that  the  liver  is  specifically  concerned  in  the 
formation  of  urea.  To  quote  the  authors :  "In  the 
absence  of  satisfactory  proof  that  deaminization  and 
urea  formation  is  localized  we  are  not  justified  in 
assuming  that  the  process  is  a  specialized  process  in 
the  sense  of  being  confined  to  some  particular  organ. 
In  other  words,  so  far  as  we  yet  know,  the  urea  form- 
ing process  is  a  characteristic  of  all  the  tissues  and  by 
far  the  greatest  amount  of  urea  is  therefore  prob- 
ably formed  in  the  muscles.  The  negative  results, 
so  far  as  any  localized  urea  formation  is  concerned,  is 
almost  satisfactory  proof  that  there  is  none,  for  if 
there  were  one  central  focus  from  which  all  or  nearly 
all  of  the  urea  originated  we  could  scarcely  have 
failed  to  find  it." 


PROTEIN  AND  CIRCULATION  73 

Are  Amino  Acids  Found  in  the  Blood? 

Opposed  to  the  investigators  advancing  the  regenera- 
tion of  protein  immediately  after  absorption  is  a 
second  group  of  men  who  have  long  believed  that 
amino  acids  are  absorbed  into  the  blood.  The  great 
difficulty  has  been  to  demonstrate  their  presence.  A 
large  number  of  experiments  have  been  devised  in 
various  ingenious  ways  to  overcome  the  difficulties 
attendant  upon  such  a  procedure.  Many  investigators 
have  obtained  partial  evidence  of  the  presence  in  the 
blood  stream  but  an  actual  isolation  and  identification 
of  individual  amino  acids  was  for  a  long  time  lacking. 

The  failure  to  obtain  definite  proof  of  the  amino 
acids  in  the  blood  has  been  due  in  large  measure  to 
the  inadequacy  of  the  methods  available.  At  any  one 
moment  the  quantity  of  these  substances  in  a  deter- 
mined sample  of  blood  must  be  exceedingly  small. 

Moreover,  one  must  remember  that  the  formation  of 
amino  acids  in  the  intestinal  tract  is  a  gradual  process 
and  not  of  the  nature  of  an  explosion  so  that  the 
quantity  of  amino  acids  available  for  passage  into  the 
blood  during  a  given  period  must  be  relatively  small. 
The  rapidity  of  circulation  is  another  factor  to  be 
taken  into  consideration.  It  has  been  shown  that  in 
the  portal  vein  of  the  dog  the  blood  travels  at  the  rate 
of  about  150  cc.  per  minute.  Pfliiger  has  estimated 
that  for  human  beings  a  maximum  rate  of  absorption 
of  protein  may  be  represented  at  1.14  gram  protein 
per  kilo  per  hour.    If  the  1.14  gram  protein  absorbed 


74  THE  AMINO  ACIDS 

per  hour  is  compared  to  the  volume  of  blood  in  which 
it  would  necessarily  be  dissolved  we  find  that  the 
protein  would  be  present  in  the  concentration  of  0.12 
per  cent.  Another  difficulty  arises  from  the  fact  that 
the  blood  is  a  fluid  already  containing  about  3  per 
cent  of  coagulable  protein  and  also  nitrogen  to  a 
smaller  extent  in  other  forms. 

Certain  fairly  definite  indications,  however,  have  led 
many  physiologists  to  maintain  a  belief  that  protein 
is  absorbed  and  is  taken  up  by  the  cells  in  the  form  of 
amino  acids. 

Determination  of  the  "residual  nitrogen"  of  the 
blood,  that  is,  the  difference  between  the  total  nitrogen 
and  that  representing  coagulable  protein  has  shown 
that  after  meals  there  is  a  slight  gain  both  in  the  portal 
blood  and  that  of  the  systematic  circulation.  This 
increase  of  nitrogen  may  be  attributed  to  the  absorbed 
amino  acids  or  polypeptides,  but  in  view  of  the  possi- 
ble existence  of  non-coagulable  proteins  in  the  blood 
it  cannot  be  accepted  as  proof  positive.  A  second 
method  for  the  same  endeavor  has  been  the  formation 
of  amino  acid  compounds  by  use  of  j8-naphthalene 
sulphochloride.  Shaken  with  the  fluid  obtained  after 
separation  of  the  serum  proteins  either  by  coagulation 
or  by  means  of  dialysis  precipitates  have  been  obtained 
with  this  reagent,  strongly  indicating  the  presence  of 
amino  acids  but  the  failure  of  the  precipitate  to  assume 
a  crystalline  form  has  made  impossible  a  positive 
identification  of  amino  acids.  Cohnheim  by  observa- 
tions with  the  isolated  intestine  of  the  octopus  was  able 


PROTEIN  AND  CIRCULATION  75 

to  prove  absorption  and  the  existence  in  the  blood  of 
certain  amino  acids  but  failed  to  detect  these  sub- 
stances when  experiments  were  carried  out  on  the 
intact  animal. 

By  the  elaboration  of  new  methods,  Folin  and  Denis 
and  Van  Slyke  and  Meyer  have  been  able  to  prove  the 
entrance  of  amino  acids  into  the  blood  stream.  Later, 
Abderhalden,  the  chief  opponent  of  the  idea  of  amino 
acid  absorption,  was  successful  in  isolating  from  the 
blood  several  of  the  individual  amino  acids  by  the 
employment  of  great  volumes  of  blood.  The  absorp- 
tion of  protein  in  the  form  of  amino  acids  having  thus 
been  established  the  question  next  arises  what  becomes 
of  them?  It  was  soon  proved  that  there  was  a  rapid 
disappearance  of  amino  acids  from  the  circulation  and 
this  fact  made  pertinent  the  queries  :  "Are  they  decom- 
posed in  the  blood:  are  they  chemically  incorporated 
into  the  complex  molecules  of  the  tissue  proteins ;  or 
are  they  merely  absorbed  by  the  tissues  in  general,  or 
by  certain  tissues  in  particular,  without  undergoing 
any  immediate  change?"  These  questions  have  been 
fully  answered  by  Van  Slyke  and  Meyer  in  experi- 
ments designed  to  follow  the  fate  of  the  amino  acids 
after  absorption.  It  was  found  that  the  amino  acids 
are  absorbed  by  the  tissues  without  undergoing  any 
immediate  chemical  change.  This  absorption  though 
rapid  is  never  complete,  the  blood  always  containing 
a  small  quantity  of  amino  acids.  It  would  appear 
from  this  that  there  is  an  equilibrium  between  the 
amino  acids  of  the  blood  and  of  the  tissues.    The  way 


76  THE  AMINO  ACIDS 

in  which  amino  acids  are  taken  up  by  the  tissues  and 
held  by  them  is  still  undetermined. 

In  a  later  communication  the  same  investigators  have 
attempted  to  determine  the  fate  of  amino  acids  after 
absorption  by  the  tissues  and  selected  the  changes 
occurring  in  the  liver.  Amino  acids  absorbed  by  the 
liver  rapidly  disappear.  In  explanation  of  this  obser- 
vation several  possibilities  exist:  1.  The  amino  acids 
may  be  excreted  through  the  bile.  This  view,  however, 
is  not  probable  since  the  quantities  of  amino  acids  in 
the  bile  and  urine  were  entirely  too  small  to  account 
for  the  amount  that  disappeared  from  the  liver.  2.  A 
second  possibility  is  that  the  amino  acids  are  trans- 
ferred to  other  tissues.  This  hypothesis  is  also  highly 
improbable  since  none  of  the  other  large  organs  show 
a  greater  avidity  for  amino  acids,  yet  three  or  four 
hours  after  injection  of  amino  acids  other  organs 
usually  contain  more  amino  acids  than  the  liver.  3. 
The  absorbed  amino  acids  are  synthesized  into  body 
protein  in  the  liver.  The  possibility  cannot  be  defi- 
nitely decided  at  present.  4.  The  amino  acids  are 
deaminated  with  formation  of  urea  or  ammonia.  In 
all  probability  a  portion  of  the  amino  acids  which 
disappears  from  the  liver  reappears  in  the  urine  as 
urea. 

The  disappearance  of  amino  acids  from  the  liver  is 
more  rapid  and  complete  than  is  true  for  other  tissues 
like  the  kidney,  intestine,  pancreas,  and  spleen.  From 
the  muscles  the  amino  acids  disappear  very  slowly. 
As  a  summary  of  the  whole  question  one  may  quote  the 


PROTEIN  AND  CIRCULATION  77 

words  of  Van  Slyke  and  Meyer:  "The  amino  adds, 
with  perhaps  some  peptides,  from  the  intestine  enter 
the  circulation,  from  which  they  are  immediately 
absorbed  by  the  tissues.  The  power  to  take  them  up 
from  the  blood  stream  is  common  to  all  the  tissues, 
but  is  limited.  The  muscles  of  the  dog,  for  example, 
reach  the  saturation  point  when  they  contain  about 
75  mgm.  of  amino  acid  nitrogen  per  100  grams.  The 
liver,  however,  continually  desaturates  itself  by  metab- 
olizing the  amino  acids  that  it  has  absorbed,  and  con- 
sequently maintains  indefinitely  its  power  to  continue 
removing  them  from  the  circulation  so  long  as  they  do 
not  enter  it  faster  than  the  liver  can  metabolize  them. 
When  the  entrance  is  unnaturally  rapid,  as  in  our 
injection  experiments,  or  when  the  liver  is  sufficiently 
degenerated,  as  observed  clinically  in  some  pathological 
conditions,  the  kidneys  assist  in  removing  the  amino 
acids  by  excreting  them  unchanged.  Death  may  result 
when  the  above  agencies  for  preventing  undue  accu- 
mulation of  protein  digestion  products  are  over-taxed. 

"In  regard  to  the  synthesis  of  tissue  proteins  it 
appears  reasonable  to  believe  that,  since  each  tissue 
has  its  o-wTi  store  of  amino  acids,  which  it  can  replenish 
from  the  blood,  it  uses  these  to  synthesize  its  own 
proteins." 

Concerning  the  manner  in  which  the  free  amino 
acids  are  utilized  by  the  tissues  two  possibilities  may 
be  assumed,  and  according  to  Van  Slyke  and  Meyer 
these  are :  1.  The  amino  acids  serve  as  a  reserve  energy 
supply,  Hke  glycogen,  or  as  a  reserve  of  tissue-building 


78  THE  AMINO  ACIDS 

material.  In  either  case  the  supply  would  be  depleted 
if  not  renewed  from  the  food.  2.  The  amino  acids 
are  merely  intermediate  steps  in  both  the  construction 
and  breakdown  of  the  tissue  proteins.  In  this  case  they 
could  originate,  not  only  from  absorbed  food  products, 
but  also  from  autolyzed  tissue  protein:  starvation 
would  not  result  in  a  disappearance  of  the  amino  acid 
supply  of  the  tissues,  and  might  even  increase  it.  To 
determine  the  correctness  of  one  or  the  other  of  these 
hypotheses  the  authors  mentioned  above  analyzed  the 
tissues  of  animals  in  various  states  of  nutrition.  The 
results  are  in  harmony  with  the  second  hypothesis,  for 
free  amino  acids  of  the  tissues  tend  to  increase  in 
starvation  rather  than  to  disappear.  The  investigators 
have  summarized  their  views  regarding  this  in  the  fol- 
lowing words :  "The  amino  acids  appear,  therefore,  to 
be  intermediate  steps,  not  only  in  the  synthesis,  but  in 
the  breaking  down  of  body  proteins.  Otherwise,  in 
order  to  explain  their  maintenance  in  the  tissues  during 
starvation,  one  would  be  forced,  contrary  to  the  con- 
clusions of  all  experimental  work  on  the  subject,  to 
assume  that  they  are  inert  substances  lying  unchanged 
for  long  periods,  even  when  most  urgently  needed  to 
build  tissue  or  supply  energy.  The  maintenance  of 
the  amino  acid  supply  by  synthesis,  from  ammonia  and 
the  products  of  fats  or  carbohydrates,  seems  excluded. 
The  supply  of  raw  material  in  the  form  of  fat  and 
carbohydrates  nearly  disappears  during  starvation,  and 
the  ammonia  could  originate  only  from  broken-down 
protein,  as  the  normal  store  of  ammonia  nitrogen  is 


PROTEIN  AND  CIRCULATION  79 

only  a  fraction  of  that  of  the  free  amino  acids.  These 
considerations,  and  the  self-evident  wasting  of  starved 
tissues,  point  strongly  to  autolysis  as  the  main  source 
of  the  free  amino  acids  in  the  fasting  body." 

"The  failure  to  increase  the  free  amino  acid  content 
of  the  tissues  by  high  protein  feeding  indicates, 
furthermore,  that  when  nitrogen  is  retained  in  the 
organism  it  is  not  to  an  appreciable  extent,  as  stored 
digestion  products,  but  rather  as  body  protein." 

These  results,  and  the  consequent  inference  from 
them,  have  made  void  all  the  older  theories  of  metab- 
olism and  it  is  becoming  more  and  more  evident  that 
in  any  consideration  of  protein  transformations  within 
the  organism  in  health  or  in  disease  amino  acids  are 
the  substances  which  demand  attention.  This  is  the 
age  of  amino  acid  metabolism  and  at  present  the  inves- 
tigations are  being  narrowed  down  to  the  point  of  the 
determination  of  what  actually  occurs  with  the  indi- 
vidual amino  acids  and  what  special  role  in  nutrition 
each  may  play. 


References  to  Literature 

Abderhalden:   Zeitschrift   fiir   physiologische   Chemie.      1913, 
88,  p.  478.    [Amino  acids  in  blood.] 

Abderhalden:  Text  Book  of  Physiological  Chemistry.     1914 
[Enzymes  in  blood.] 

Cathcart:  The  Physiology  of  Protein  Metabolism.    1912. 


80  THE  AMINO  ACIDS 

Folin  and  Denis:  Journal  of  Biological  Chemistry.  1912,  ii, 
p.  87  and  p.  161;  1912,  I2,  p.  141  and  p.  253.  [Fate  of 
digestion  products.] 

Mendel:  Theorien  des  Eiweissstoffwechsels  nebst  einigen 
praktischen  Konsequenzen  derselben.  Ergebnisse  der 
Physiologic.    1911,  n,  p.  418. 

Van  Slyke  and  Meyer:  Journal  of  Biological  Chemistry.  1912, 
12,  p.  399;  1913-1914,  i6,  p.  187,  p.  197,  p.  213,  and  p.  231. 
[Fate  of  digestion  products.] 


CHAPTER  V 

THEORIES  OF  PROTEIN  METABOLISM 

Under  the  term  protein  metabolism  are  included  all 
the  processes  in  the  animal  organism  concerned  with 
the  fate  of  protein  whether  introduced  as  food  or 
serving  as  tissue  substance.  The  metabolic  changes 
are  divisible  into  two  distinct  phases  known  as  anabo- 
lism,  or  building  up  processes,  and  cataholism,  or  de- 
structive processes.  Although  it  is  definitely  recog- 
nized that  metabolic  activity  is  manifested  in  two  dia- 
metrically opposed  directions,  the  successive  stages  in 
either  process  are  but  vaguely  understood.  Only  the 
starting  point  and  the  final  end  products  in  each  in- 
stance can  be  stated  with  certainty,  although  here  and 
there  individual  stages  in  the  processes  under  discus- 
sion point  in  one  or  another  direction,  and  thus  give 
indication  of  the  type  of  activity  that  must  have  gone 
before.  For  the  unravelling  of  the  mystery  the  first 
requisite  is  a  clear  conception  of  the  problem.  In  the 
words  of  Liebig :  "If  we  take  the  letters  of  a  sentence 
which  we  wish  to  decipher,  and  place  them  in  a  line, 
we  advance  not  a  step  towards  the  discovery  of  their 
meaning.  To  resolve  an  enigma,  we  must  have  a  per- 
fectly clear  conception  of  the  problem.  There  are 
many  ways  to  the  highest  pinnacle  of  a  mountain ;  but 


82  THE  AMINO  ACIDS 

those  only  can  hope  to  reach  it  who  keep  the  summit 
constantly  in  view.  All  our  labor  and  all  our  efforts, 
if  we  strive  to  attain  it  through  a  morass,  only  serve 
to  cover  us  more  completely  with  mud;  our  progress 
is  impeded  by  difficulties  of  our  own  creation,  and  at 
last  even  the  greatest  strength  must  give  way  when  so 
absurdly  wasted." 

The  development  of  knowledge  in  science  succeeds 
best  when  an  hypothesis  is  formulated  as  a  basis  for 
investigation.  By  holding  fast  to  that  which  is  proven 
as  fact  and  discarding  that  which  is  shown  to  be  con- 
trary to  fact  is  real  progress  made.  This,  indeed,  has 
been  the  case  in  the  history  of  protein  metabolism,  as 
may  be  seen  in  the  following  pages  where  is  traced  the 
evolution  of  ideas  concerning  it.  . 

LlEBIG 

The  first  clearly  defined  theory  of  protein  metabo- 
lism was  that  enunciated  by  Liebig  who  assumed  that 
protein  material  undergoes  little  or  no  chemical  change 
previous  to  its  introduction  into  the  blood  stream  and 
its  assimilation  by  the  tissues.  "According  to  this 
theory,  the  plant  holds  a  position  intermediate  between 
the  mineral  and  animal  world.  The  animal  is  incapable 
of  assimilating  the  compounds  stored  up  in  inorganic 
nature.  To  render  these  compounds  subservient  to 
the  purposes  of  animal  life  they  may  have  to  undergo 
a  preliminary  preparation  within  the  living  organism  of 
the  plant.    The  simple  mineral  molecules  are  thus  con- 


PROTEIN  METABOLISM  83 

verted  into  molecules  of  a  higher  order,  fit  to  serve  in 
building  up  and  maintaining  alive  the  body  of  the 
animal."  "How  admirably  simple  after  we  have 
acquired  a  knowledge  of  this  relation  between  plants 
and  animals,  appears  to  us  the  process  of  formation 
of  the  animal  body,  the  origin  of  its  blood  and  organs ! 
The  vegetable  substances,  which  serve  for  the  produc- 
tion of  blood,  contain  already  the  chief  constituent  of 
blood  ready  formed,  with  all  its  elements."  "The  true 
starting  point  for  all  the  tissues  is,  consequently, 
albumen ;  all  nitrogenized  articles  of  food,  whether 
derived  from  animal  or  from  the  vegetable  kingdom, 
are  converted  into  albumen  before  they  can  take  part 
in  the  process  of  nutrition." 

According  to  Liebig  digestion  is  merely  a  process 
whereby  food  becomes  changed  to  a  soluble  condition 
capable  of  absorption  without  transformation  of  its 
identity.  This  soluble  albumen  is  built  up  into  organ- 
ized tissue  previous  to  its  degradation  (an  idea  later 
adopted  by  Pfliiger).  Thus  we  read:  "There  can  be 
no  greater  contradiction,  with  regard  to  the  nutritive 
process,  than  to  suppose  that  the  nitrogen  of  the  food 
can  pass  into  the  urine  as  urea,  without  having  pre- 
viously become  part  of  an  organized  tissue;  for  albu- 
men, the  only  constituent  of  blood  which,  from  its 
amount,  ought  to  be  taken  into  consideration,  suffers 
not  the  slightest  change  in  passing  through  the  liver  or 
kidneys ;  we  find  it  in  every  part  of  the  body  with  the 
same  appearance  and  the  same  properties." 

Liebig  divided  all  foods  into  two  groups,  the  nitro- 


84:  THE  AMINO  ACIDS 

genous,  or  plastic,  foods,  and  the  non-nitrogenous  or 
respiratory  foods.  In  accordance  with  this  classifica- 
tion plastic  foods  were  tissue  formers  and  supplied 
energy  for  muscular  activity;  the  respiratory  foods, 
on  the  other  hand,  were  essential  for  the  respiratory 
act  and  the  constant  temperature  of  the  body,  but 
could  not  be  transformed  into  organized  tissue. 

VoiT 

The  fundamental  conception  of  Voit  (1867)  was 
that  all  protein  in  the  body  is  not  decomposed  with 
equal  ease.  In  accordance  with  this  idea  he  divided 
the  protein  material  of  the  body  into  two  groups,  the 
organised  or  tissue  protein,  that  built  up  into  living 
protoplasm  and  difficult  of  disintegration  and,  secondly, 
circulating  protein  existing  in  the  fluids  and  tissues  of 
the  organism  without  being  an  integral  part.  The 
circulating  protein  may  be  more  easily  and  readily 
destroyed  than  the  organized  or  tissue  protein.  In  his 
classic  experiment  designed  to  show  the  difference  of 
metabolism  between  tissue  protein  and  circulating 
protein,  Voit  allowed  a  well-fed  dog  to  starve  for 
several  days.  He  demonstrated  that  under  these  cir- 
cumstances there  is  at  first  an  abundant  decomposition 
of  protein  material  which  is  later  followed  by  a  period 
during  which  very  little  protein  is  catabolized.  His 
interpretation  of  these  facts  was  to  the  effect  that  dur- 
ing the  time  when  a  large  protein  disintegration  ob- 
tained only  circulating  protein  was  destroyed,  whereas 


PROTEIN  METABOLISM  85 

in  the  later  stages  the  greatly  diminished  uniform 
protein  metabolism  was  that  of  the  organized  or  tissue 
proteins. 

In  his  theory  Voit  assigned  to  the  cells  the  function 
pf  utilizing  proteins,  the  older  view  that  metabolism 
took  place  in  the  blood  having  been  discarded.    From 
the  fluids  bathing  the  tissues  food  or  circulating  pro- 
■(ein  is  drawn  within  the  cells  and  there  transformed. 
On  the  other  hand,  a  certain  small  amount  of  tissue 
protein  is  constantly  dying  and  is  replaced  by  circulat- 
ing protein,  thus  becoming  eventually  living  proto- 
plasm.    "The  tissue-elements  constitute  an  apparatus 
of  a  relatively  stable  nature,  which  has  the  power  of 
taking  proteins  from  the  fluids  washing  the  tissues  and 
appropriating  them,  while  their  own  proteins,  the  tissue 
proteins,  are  ordinarily  catabolized  to  only  a  small  ex- 
tent, about  1  per  cent  daily."     (Voit.)     "By  an  in- 
creased supply  of  proteins  the  activity  of  the  cells  and 
their  ability  to  decompose  nutritive  proteins  are  also 
increased   to   a   certain   degree.     When   nitrogenous 
equilibrium  is  obtained  after  an  increased  supply  of 
proteins,  it  indicates  that  the  decomposing  power  of 
the  cells  for  proteins  has  increased  so  that  the  same 
quantity  of  proteins  is  metabolized  as  is  supplied  to  the 
body.     If  the  protein  metabolism  is  decreased  by  the 
simultaneous  administration  of  other  non-nitrogenous 
foods,  a  part  of  the  circulating  proteins  may  have  time 
to  become  fixed  and  organized  by  the  tissues,  and  in 
this  way  the  flesh  of  the  body  increases.     During 
starvation  or  with  a  lack  of  protein  in  the  food  the 


86  THE  AMINO  ACIDS 

reverse  takes  place,  for  a  part  of  the  tissue  proteins  is 
converted  into  circulating  proteins,  which  are  metabo- 
lized, and  in  this  case  the  flesh  of  the  body  decreases." 
(Hammarsten.) 

Pfluger 

In  1893  Pfluger  severely  criticised  the  theory  of 
Voit  and  offered  another  in  its  place.  In  its  essence 
the  theory  of  Pfliiger  rests  upon  the  hypothesis  that 
food  protein  must  become  living  protoplasm  before 
it  can  be  utilized  for  the  needs  of  the  body.  In  accord- 
ance with  this  idea  he  assumed  that  food  protein  is 
catabolized  with  great  difficulty  whereas  living  proto- 
plasm is  in  a  state  of  continual  unstable  equilibrium 
leading  to  any  easy  oxidation  or  decomposition  of  its 
protein.  Pfliiger's  theory  rests  upon  experiments 
carried  through  by  his  pupil  Schondorff .  It  was  shown 
by  Schondorff  that  when  the  blood  from  a  starving 
dog  was  passed  through  the  hind  limbs  and  liver  of 
a  well-fed  animal  the  urea  of  this  blood  was  increased. 
On  the  other  hand,  no  increase  of  urea  could  be  ob- 
tained when  blood,  whether  of  starved  or  well-fed 
dogs,  was  passed  through  the  hind  limbs  and  liver  of 
a  starved  dog.  From  the  results  Pfliiger  argued  that 
the  determining  factor  in  protein  catabolism  is  the 
state  of  nutrition  in  the  tissue  cells  and  not  the  cir- 
culating protein. 

Although  in  general  Pfliiger  appeared  to  disprove 
many  of  the  points  in  Voit's  theory,  one  positive  evi- 
dence stands  out  clearly  in  favor  of  Voit's  theory,  and 


PROTEIN  METABOLISM  87 

that  is  the  fact  of  the  rapidity  with  which  large  quan- 
tities of  protein  are  cataboHzed  in  the  body.  In  a  few 
hours  great  quantities  of  protein  may  be  disintegrated 
as  judged  by  the  corresponding  increase  in  urinary 
nitrogen.  It  is  hardly  probable  that  living  protoplasm 
could  be  synthesized  so  rapidly  and  so  much  of  it  be 
so  quickly  destroyed  again.  This  is  the  more  incredi- 
ble since  the  same  fact  applies  irrespective  of  the 
previous  state  of  nutrition  of  the  organism. 

In  1905  Folin  subjected  the  experiments  of  Schon- 
dorff  to  a  searching  criticism  and  pointed  out  that  the 
evidence  furnished  by  them  was  by  no  means  unassail- 
able. Upon  studying  the  details  of  one  of  Schondorif's 
experiments,  Folin  found  that  the  actual  increase  in 
urea  nitrogen  in  the  transfused  blood  amounted  to  less 
than  one-tenth  of  1  per  cent  instead  of  125  per  cent  as 
calculated  by  Schondorff.  "Considering  the  numerous 
sources  of  error  and  uncertainty  necessarily  attached 
to  an  experiment  of  this  kind,  it  seems  very  strange 
that  the  extraction  of  25  mgm.  of  urea-nitrogen  from 
the  hind  limbs  of  a  dog  killed  while  engaged  in  digest- 
ing 700  gm.  of  meat  should  be  accepted  as  proving  not 
only  that  protein  catabolism  did  occur  during  the 
experiment,  but  also  that  it  occurred  in  the  bioplasm 
and  not  in  the  circulating  protein." 

No  direct  evidence  has  been  obtained  to  prove  or  dis- 
prove the  one  or  the  other  of  these  last  two  widely 
divergent  theories.  The  distinction  between  tissue 
protein  and  food  protein  is  probably  one  of  degree 
rather  than  of  kind. 


88  THE  AMINO  ACIDS 

ICASSOWI.TZ 

Kassowitz  in  1904  put  forth  the  view  that  it  was 
scarcely  probable  that  a  substance  would  serve  both  as 
reconstructive  material  for  disintegrated  cells  and  as 
a  source  of  energy.  According  to  his  ideas  food  pro- 
tein is  not  merely  transformed  into  living  protoplasm 
by  some  obscure  rearrangement  but  there  is  an  actual 
synthesis  with  fat  and  carbohydrate  to  form  living 
bioplasm.  Like  Pfiiiger  he  adopts  the  view  that  only 
^'organized"  protein  is  oxidized. 

In  metabolism  there  are  two  types  of  protoplasmic 
disintegration:  the  inactive,  whereby  the  protoplasm 
formed  from  food  protein  during  rest  is  immediately 
changed  or  broken  down  into  non-nitrogenous  storage 
materials  (glycogen  and  fat)  and  urea;  the  active,  by 
which  under  the  influence  of  stimuli  which  induce 
muscular  contractions,  the  protein  nucleus  of  the  dis- 
integrating protoplasm  molecule  is  left  intact  so  that 
it  may  serve  for  the  resynthesis  of  protoplasm  with 
fresh  non-nitrogenous  compounds.     (Mendel.) 

Speck 

In  the  theory  of  Speck  (1903)  the  view  is  held  that 
two  forms  of  protein  exist  but  that  the  catabolism  of 
organized  protein  is  quite  different  from  that  of  the 
unorganized  protein.  That  portion  of  food  protein 
(unorganized)  not  employed  for  the  building  up  of 
living  tissue,  is  split  into  two  portions,  first,  a  nitro- 
genous part,   which  is  rapidly  converted  into   urea, 


PROTEIN  METABOLISM  89 

and  a  nitrogen  free  residue,  serving  as  a  ready  source 
of  energy. 

On  the  other  hand,  after  death  of  cells,  the  tissue 
or  organized  protein,  although  also  broken  down  into 
two  parts,  finds  a  destiny  unlike  the  products  of  food 
protein  disintegration.  Tissue  protein  splits  into  a 
nitrogen-containing  and  a  nitrogen-free  portion.  Under 
normal  conditions  the  nitrogen-free  part  is  trans- 
formed to  glycogen  or  fat  which  may  be  utilized  for 
purposes  of  energy.  The  portion  containing  nitrogen 
is  not  broken  down  at  once  into  urea  but  it  leads  to 
the  formation  of  a  variety  of  substances  which  play 
important  roles  in  metabolism  but  are  finally  excreted 
as  urea.  In  the  decomposition  of  tissue  protein  Speck 
assigned  to  oxygen  deficiency  an  exceedingly  important 
part. 

RUBNER 

In  Rubner's  theory  of  protein  metabolism  it  is  main- 
tained that  a  study  of  metabolism  cannot  be  considered 
separately  from  the  study  of  heat  production.  Ac- 
cording to  Rubner,  therefore,  metabolism  must  be 
studied  in  connection  with  the  exchange  of  energy. 
In  all  of  the  metabolic  changes  undergone  by  protein 
in  the  body  reference  is  made  to  the  accompanying 
production  of  heat.  Rubner  believes  in  a  "store"  pro- 
tein which  may  be  compared  to  Voit's  "circulating" 
protein,  and  in  a  "wear  and  tear  quota"  necessary  for 
the  repair  of  tissue  waste.  He  assumes  that  most  of 
the  protein  after  absorption  is  rapidly  split  into  two 


90  THE  AMINO  ACIDS 

parts,  one  nitrogen-free,  the  other  containing  nitrogen. 
Inasmuch  as  the  nitrogen-containing  part  plays  Httle 
role  in  energy  exchange,  its  fate  is  left  somewhat  in- 
definite. The  part  free  from  nitrogen  forms  the 
dynamic  quota  of  the  protein  ingested.  When  protein 
is  disintegrated  into  its  two  parts  mentioned  above, 
there  occurs  a  certain  liberation  of  heat  which  is  of  no 
value  to  the  body  cells  and  is  therefore  lost.  This  lib- 
eration of  energy  has  been  called  by  Rubner  the  "spe- 
cific dynamic  action"  of  protein. 

"A  highly  speculative  hypothesis  explained  how  the 
various  changes  took  place.  All  protoplasm  was  not 
regarded  as  being  of  the  same  type,  one  kind  might 
be  thermolabile,  another  thermostable,  but  all  varieties 
had  in  common  a  certain  molecular  grouping  which 
acted  as  a  kind  of  nucleus  to  which  other  protein 
groups  (for  example  those  which  were  thermostable 
or  thermolabile)  could  attach  themselves.  The  mech- 
anism of  the  energy  exchange,  which  is  characteris- 
tic of  activity,  was  effected  by  a  distinct  vibratory 
movement  of  the  whole  or  a  definite  part  of  the  proto- 
plasm. Owing  to  the  specific  oscillation,  the  proto- 
plasm had  the  power  of  bringing  about  the  brerl^'iown 
of  contiguous  foodstuffs.  The  'affinities'  (specific 
oscillations)  must  be  of  a  specific  nature  for  each 
tissue  and  were  probably  somewhat  akin  to  ferment 
action.  Thus,  in  diabetes,  the  'affinities'  ♦rhich 
brought  about  the  breakdown  of  carbohydrates,  were 
for  some  reason  or  other  in  a  state  of  suspension, 
inoperative  or  actually  destroyed,  whereas  those  which 


PROTEIN  METABOLISM  91 

dealt  with  the  catabolism  of  fat  were  active.  The 
direct  effect  of  the  approximation  of  the  foodstuffs 
to  the  'affinities'  resulted  in  an  atomic  rearrangement 
and  the  entry  of  oxygen.  The  potential  energy  of  the 
foodstuff  now  became  available  and  caused  a  complete 
alteration  in  the  'affinities';  an  absorption  of  energy 
into  the  living  substance  took  place  at  the  moment  of 
the  catabolism  of  the  foodstuff.  The  internal  oscilla- 
tions and  changes  in  the  cells,  however,  gradually  used 
up  all  the  energy,  which  was  converted  into  heat  and 
lost,  and  there  was  a  return  to  the  original  condition, 
the  'affinities'  being  again  ready  to  begin  work.  The 
rate  of  the  change  depended  on  the  nature  of  the 
living  substance,  the  temperature,  nervous  influences, 
and  the  conditions  of  the  organism  itself."   (Cathcart.) 

FOLIN 

It  was  Folin's  conception  that  "the  laws  governing 
the  composition  of  the  urine  represent  only  the  effects 
of  other  more  important  laws  governing  the  catabo- 
lism of  protein  in  the  animal  organism"  which  led 
him  to  determine  these  laws  under  widely  differing 
conditions  of  diet.  His  interpretation  of  protein 
metabolism  on  the  basis  of  observed  variations  in  the 
percentage  composition  of  the  urine  has  stood  as  the 
almost  universally  accepted  theory  of  protein  metabo- 
lism of  the  present  period. 

Previous  to  his  investigation  only  lengthy  and  none 
too  accurate  methods  were  in  use  for  the  estimation 


92 


THE  AMINO  ACIDS 


of  the  urinary  constituents.  He,  therefore,  first  de- 
vised a  method,  in  each  instance  short  and  accurate, 
for  the  estimation  of  every  important  nitrogenous 
constituent  of  the  urine,  together  with  methods  for  the 
determination  of  sulphur  containing  compounds,  and, 
secondly,  with  the  aid  of  these  methods  made  complete 
analysis  of  normal  urines.  In  order  to  make  the  factor 
of  food  protein  as  evident  as  possible,  diets  rich  in 
protein  were  fed  and  were  succeeded  by  rations 
markedly  deficient  in  nitrogenous  substances  although 
containing  a  sufficiency  of  energy  yielding  substances. 
As  showing  the  wide  range  of  variation  on  the  two 
diets  a  typical  example  of  the  urinary  composition 
follows : 

Nitrogen  rich  diet      Nitrogen  poor  diet 

1170  c.c.  38Sc.c. 

16.8  grams  3.60  grams 

14.7  grams  =  87.5%  2.20  grams  =  61.7% 

0.49  gram   =   3.0%  0.42  gram  =11.3% 

0.18  gram   =    1.1%  0.09  gram   =   2.5% 

0.58  gram  =   3.6%  0.60  gram   =17.2% 


Volume  of  urine 

Total  Nitrogen 

Urea-Nitrogen 

Ammonia-Nitrogen 

Uric  acid-Nitrogen 

Kreatinine-Nitrogen 

Undetermined 

Nitrogen 
Total  S03 
Inorganic  SO^ 
Ethereal  SO^ 
Neutral  SO^ 


0.85  gram   =   4.9% 
3.64  grams 
3.27  grams 
0.19  gram 
0.18  gram 


90.0% 

5.2% 
4.8% 


0.27  gram 
0.76  gram 
0.46  gram 
0.10  gram 
0.20  gram 


=   7.3% 

=  60.5% 
=  13.2% 
=  26.3% 


The  general  laws  deduced  by  Folin  as  a  result  of 
urinary  analysis  are : 

1.  Kreatinine.  The  absolute  quantity  of  kreatinine 
eliminated  on  a  meat-free  diet  is  a  constant  quantity, 


PROTEIN  METABOLISM  93 

different  for  different  individuals,  but  wholly  inde- 
pendent of  quantitative  changes  in  the  total  amount  of 
nitrogen  eliminated. 

2.  Uric  Acid.  When  the  total  amount  of  protein 
metabolism  is  greatly  reduced,  the  absolute  quantity  of 
uric  acid  is  diminished,  but  not  nearly  in  proportion  to 
the  diminution  in  the  total  nitrogen,  and  the  per  cent 
of  the  uric  acid  nitrogen  in  terms  of  the  total  is,  there- 
fore, much  increased. 

3.  Ammonia.  With  pronounced  diminution  in  the 
protein  metabolism  (as  shown  by  the  total  nitrogen  in 
the  urine),  there  is  usually,  but  not  always,  and  there- 
fore not  necessarily,  a  decrease  in  the  absolute  quan- 
tity of  ammonia  eliminated.  A  pronounced  reduction 
of  the  total  nitrogen  is,  however,  always  accompanied 
by  a  relative  increase  in  the  ammonia-nitrogen,  pro- 
vided that  the  food  is  not  such  as  to  yield  an  alkaline 
ash. 

4.  Urea.  With  every  decided  diminution  in  the 
quantity  of  total  nitrogen  eliminated,  there  is  a  pro- 
nounced reduction  in  the  per  cent  of  that  nitrogen 
represented  by  urea.  When  the  daily  total  nitrogen 
elimination  has  been  reduced  to  3  gm.  or  4  gm.  about 
60  per  cent  of  it  only  is  in  the  form  of  urea. 

5.  Inorganic  Sulphates.  Decided  diminutions  in 
the  daily  elimination  of  total  sulphur  are  accompanied 
by  reductions  in  the  per  cent  of  the  sulphur  present  as 
inorganic  sulphates.  The  reductions  are  as  great  as  in 
the  case  of  urea. 

6.  Neutral  Sulphur.    The  neutral  sulphur  elimina- 


y 


94  THE  AMINO  ACIDS 

tion  is  analogous  to  that  of  the  kreatinine.  It  repre- 
sents products  which  in  the  main  are  independent  of 
the  total  amount  of  sulphur  eliminated  or  of  protein 
catabolized. 

7.  Ethereal  Sulphates.  The  ethereal  sulphates  rep- 
resent a  form  of  sulphur  metabolism  which  becomes 
more  prominent  when  the  food  contains  little  or  no 
protein. 

Folin  concludes  that  neither  the  theory  of  Voit  nor 
that  of  Pfluger  can  be  correct  for  these  theories  do  not 
harmonize  with  the  above  laws  governing  the  compo- 
sition of  the  urine.  With  respect  to  his  own  views  he 
says:  "We  have  seen  (from  the  tables)  that  the  com- 
position of  urine,  representing  15  gm.  of  nitrogen,  or 
about  95  gm.  of  protein,  differs  very  widely  from  the 
composition  of  urine  representing  only  3  gm.  or  4 
gm.  of  nitrogen,  and  that  there  is  a  gradual  and  regu- 
lar transition  from  the  one  to  the  other.  To  explain 
such  changes  in  the  composition  of  the  urine  on  the 
basis  of  protein  catabolism,  we  are  forced,  it  seems 
to  me,  to  assume  that  catabolism  is  not  all  of  one 
kind.  There  must  be  at  least  two  kinds.  Moreover, 
from  the  nature  of  the  changes  in  the  distribution  of 
the  urinary  constituents,  it  can  be  affirmed,  I  think, 
that  the  two  forms  of  protein  catabolism  are  essentially 
independent  and  quite  different.  One  kind  is  extremely 
variable  in  quantity,  the  other  tends  to  remain  constant. 
The  one  kind  yields  chiefly  urea  and  inorganic  sul- 
phates, no  kreatinin,  and  probably  no  neutral  sulphur. 
The  other,  the  constant  catabolism,  is  largely  repre- 


PROTEIN  METABOLISM  95 

sented  by  kreatinin  and  neutral  sulphur,  and  to  a  less 
extent  by  uric  acid  and  ethereal  sulphates.  The  more 
the  total  catabolism  is  reduced,  the  less  prominent  be- 
come the  two  chief  representatives  of  the  variable 
catabolism." 

"The  fact  that  the  urea  and  inorganic  sulphates 
represent  chiefly  the  variable  catabolism  does  of  course 
not  preclude  the  possibility  that  they  also  represent  to 
some  extent  the  constant  catabolism." 

In  accordance  with  these  two  types  of  catabolism 
Folin  has  furnished  suitable  names.  The  protein 
metabolism  which  tends  to  be  constant  is  tissue  metab- 
olism, or  endogenous  metabolism ;  the  other,  the  vari- 
able protein  metabolism,  is  the  exogenous  or  inter- 
mediate metabolism. 

Instead  of  assuming,  as  did  Voit  and  Pfliiger,  that 
the  same  type  of  decomposition,  i.e.,  oxidation,  occurs 
with  protein  as  with  fats  and  carbohydrates,  Folin 
advances  the  view  that  the  disintegration  of  protein 
in  catabolism  is  produced  in  large  measure  by  a  series 
of  hydrolytic  splittings,  nitrogen  being  split  off  as 
ammonia. 

It  is  further  shown  that  contrary  to  the  ideas  of 
Voit  and  Pfliiger,  extensive  formation  of  urea  does 
not  occur  in  the  muscles.  Folin  believed  (1905)  that 
the  nitrogenous  cleavage  products  formed  in  the  ali- 
mentary canal  from  food  protein  are  denitrogenized, 
probably  in  the  intestine,  the  ammonia  split  off,  carried 
to  the  liver,  built  up  into  urea  and  eliminated.  The 
non-nitrogenous    residue   is    in   part   converted    into 


96  THE  AMINO  ACIDS 

carbohydrates.  "The  chief  reason  why  the  nitro- 
genous spUtting  products  produced  by  the  digestive 
enzymes  are  universally  assumed  to  be  reconverted 
into  albumin  is  the  teleological  one.  The  food  proteins 
are  tissue  builders  and  the  organism  must  not  waste 
them.  The  fact  that  the  muscle  tissues  of  normal  men 
do  not  increase  when  the  protein  of  food  is  increased, 
but  that  all  of  the  nitrogen  of  such  protein  is  at  once 
eliminated,  has  not  been  sufficiently  considered  in  this 
connection.  The  only  adequate  teleological  explana- 
tion of  this  fact  is  that  this  nitrogen  is  not  needed  for 
the  building  of  new  tissues.  It  is  not  needed  because 
the  organism  cannot  enlarge  indefinitely,  and  because 
after  it  has  attained  its  full  growth  the  daily  waste  of 
tissue  is  small.  Yet  when  more  nitrogen  than  the 
organism  needs  is  furnished  with  the  food,  we  find 
that  the  protein  containing  it  is  still  absorbed  up  to 
the  limit  of  the  digestive  capacity."  "The  greater  part 
of  the  protein  furnished  with  standard  diets  like  Voit's, 
i.e.,  that  part  representing  the  exogenous  metabolism, 
is  not  needed,  or  to  be  more  specific,  its  nitrogen  is  not 
needed.  The  organism  has  developed  special  facilities 
for  getting  rid  of  such  excess  of  nitrogen  so  as  to  get 
the  use  of  the  carbonaceous  part  of  the  protein  con- 
taining it.  The  first  step  in  this  process  is  the  decom- 
position of  protein  in  the  digestive  tract  into  proteoses, 
amido  acids,  ammonia,  and  possibly  urea.  The  hydro- 
lytic  decompositions  are  carried  further  in  the  mucous 
membrane  of  the  intestines,  and  are  completed  in  the 


PROTEIN  METABOLISM  97 

liver,  each  splitting  being  such  as  to  further  the  forma- 
tion of  urea." 

"In  these  special  hydrolytic  decompositions,  the 
result  of  which  is  to  remove  the  unnecessary  nitrogen, 
we  have  an  explanation  of  why  and  how  the  animal 
organism  tends  to  maintain  nitrogen  equilibrium  even 
when  excessive  amounts  of  protein  are  furnished  with 
the  food.  This  excess  of  protein  is  not  stored  up  in 
the  organism,  as  such,  because  the  actual  need  of  nitro- 
gen is  so  small  that  an  excess  is  always  furnished  with 
the  food.  ..." 

Present-Day  Theory  of  Metabolism 

The  proof  of  the  presence  of  amino  acids  in  the 
blood  through  the  investigations  of  Folin,  Van  Slyke, 
and  others  has  rendered  necessary  some  slight  modifi- 
cation of  our  views  concerning  metabolic  processes. 
Although  Folin  in  his  original  theory  foretold  the 
probable  importance  of  the  lower  protein  decomposi- 
tion products  it  was  not  until  the  actual  presence  of 
these  substances  in  the  blood  and  tissues  was  demon- 
strated that  acceptance  of  this  idea  was  general.  Since 
it  has  been  proven  beyond  question  that  amino  acids 
are  normally  absorbed  directly  into  the  blood  from  the 
intestine  and  are  distributed  to  the  tissues,  it  is  assumed 
that  each  tissue  rebuilds  itself  from  the  mixture  of 
amino  acids  thus  received.  That  portion  of  amino 
acids  which  is  not  necessary  for  synthesis  is  changed 
into  urea  and  carbonaceous  residues  presumably  by  a 


98  THE  AMINO  ACIDS 

process  of  deamination.  The  carbon  remainders  may- 
be transformed  into  carbohydrate  or  in  other  ways 
changed  so  as  to  yield  energy  and  heat.  Protein 
material  broken  down  within  the  tissues  undoubtedly 
suffers  a  series  of  hydrolytic  cleavages,  resulting  in  the 
formation  of  amino  acids  and  the  latter  presumably 
undergo  the  same  fate  as  those  produced  from  food 
protein. 

According  to  this  view  protein  synthesis  is  not 
restricted  to  any  one  organ  or  tissue  but  all  possess  the 
same  property.  Urea  formation  also  can  no  longer 
be  assigned  to  the  liver  or  some  special  urea-forming 
organ,  but  on  the  other  hand  every  tissue  probably  is 
capable  of  forming  this  substance. 

References  to  Literature 

Cathcart:  The  Physiology  of  Protein  Metabolism,    1912. 

Folin:  American  Journal  of  Physiology.    1905,  13,  p.  45. 

Folin:  Intermediary  Protein  Metabolism.  Journal  of  the 
American  Medical  Association.    1914,  63 j  p.  823. 

Hammarsten:  Text  Book  of  Physiological  Chemistry.     1914. 

Liebig:  Complete  Works  on  Chemistry.     1856. 

Mendel:  Theorien  des  Eiweissstoffwechsels  nebst  einigen 
praktischen  Konsequenzen  derselben.  Ergebnisse  der 
Physiologic.    1911,  I  J,  p.  418. 


CHAPTER  VI 

THE  FURTHER  FATE  OF  AMINO  ACIDS 

It  is  well  recognized  that  a  large  part  of  the  amino 
acids  of  the  food  is  eliminated  from  the  body  in  the 
form  of  urea,  carbon  dioxide,  and  water.  The  various 
amino  acids  presumably  undergo  a  variety  of  chemical 
changes  previous  to  their  excretion  as  the  simple  pro- 
ducts mentioned  above.  Also  the  ease  with  which  these 
transformations  take  place  is  different  for  the  indi- 
vidual amino  acids.  The  steps  leading  to  the  ultimate 
fate  of  some  are  quite  clear,  of  others  it  is  very  ob- 
scure or  entirely  unknown.  The  unlike  ease  of  trans- 
formation of  amino  acids  into  urea  has  been  shown 
by  intravenous  injection.  Thus  glycocoll  and  leucine 
yield  urea  more  or  less  completely  whereas  alanine, 
cystine,  aspartic  and  glutamic  acids  are  not  readily 
catabolized. 

In  general  the  first  step  in  the  metabolism  of  amino 
acids  is  that  of  oxidative  deamination — a  splitting  off 
of  ammonia  with  an  accompanying  oxidation.  For 
any  straight  chain  amino  acid  the  reaction  occurring 
may  be  represented  as  follows : 

R.CH2.CH.NH2.COOH  -f-  O  = 

R.CH,.COOH  +  CO,  +  NH3 


100  THE  AMINO  ACIDS 

The  CO2  and  urea  are  then  synthesized  to  form  urea. 
This  synthesis  may  occur  according  to  our  present 
views  in  any  active  tissue  or  organ.  Taking  leucine  as 
a  specific  example  of  oxidative  deamination  we  have 
the  reaction  following: 


CxioCH*  CHaCHj 


CH  CH 

I  I 

CH,  +  O,  CH,  +  CO,  +  NH, 

CH.NH,  COOH 

COOH 
Leucine  Isovaleric  acid 

It  has  been  shown  that  under  suitable  conditions  leu- 
cine, for  example,  may  yield  acetone.  In  order  to 
explain  the  chemistry  of  this  change  it  becomes  neces- 
sary to  introduce  the  intervention  of  a  type  of  acid 
known  as  a  ketone  acid,  that  is,  one  possessing  the 
ketone  group,  C  =:  O.  Leucine  by  oxidative  deamina- 
tion may  be  changed  to  a  ketone  acid. 


CH3CH3 

CH3CH, 

\/ 

\/ 

CH 

by  oxidative 

CH 

CH, 

deamination 
becomes 

CH, 

CH.NH, 

c  =  0 

COOH 

COOH 

Leucine 

Ketone  acid 

FURTHER  FATE  OF  AMINO  ACIDS    101 

The  ketone  acid  is  then  transformed  to  a  lower  fatty- 
acid,  isovaleric  acid,  by  cleavage  of  CO2. 


CH3CH,  CH3CH3  CH, 


)8  CH  ,  C  =  O  COOH 

I  by 

cleavage 

I  ^  becomes 

a  CHa 

COOH 

Isovaleric  acid  Acetone  Acetic  acid 

By  cleavagfe  of  isovaleric  acid  between  the  a  +  ^ 
carbon  atoms  acetone  and  acetic  acid  may  be  formed, 
both  of  w^hich  may  finally  yield  COg  +  HgO. 

Another  possibility  of  the  transformation  of  straight 
chain  amino  acids  is  first  the  formation  of  hydroxy- 
amino  acids,  then  oxidative  deamination  with  the  sub- 
sequent splitting  off  of  CO2  from  the  nitrogen  free 
residue,  or  fatty  acid,  and  the  final  direct  change  of  the 
latter  to  CO2+H2O,  thus : 

R.CH2.CH.NH2.COOH  is  first  changed  to 
R.CH2.C(OH).NH2.COOH 
Hydroxy-amino  acid 

then  oxidative  deamination  follows  with  the  formation 
of  a  ketone  acid. 


102  THE  AMINO  ACIDS 

R.CH2.C(OH).NH2.COOH 
Hydroxy-amino  acid 
R .  CH2 .  CO .  COOH  +  CO2  +  NH3 
Ketone  acid 

By  cleavage  of  COg  this  becomes  a  fatty  acid  with  less 
carbon  atoms. 

R. CH2. CO. COOH  — COgrrrR.CHg. COOH 
"Ketone  acid 

By   further   cleavage   this    fatty   acid   is   changed   to 
CO2  +  H^O. 

In  an  abnormal  organism,  such  as  that  of  the  dia- 
betic, leucine  may  yield  beta-oxybutyric  acid  instead  of 
acetone.    The  reactions  involved  follow. 


CH3CH3 

CH 

CH3CH3 

CH 

\ 

CH3CH3 
\/ 

CH 
_\                 

CH3 
CHOH 

CH, 

'^     CH, 

^    CH, 

^    CH, 

CH.NH, 

CHOH 

COOH 

COOH 

COOH 

COOH 

Leucine 

Oxy-isobutyl 
acetic  acid 

Isovaleric 
acid 

jS-oxy-butyric 
acid 

FURTHER  FATE  OF  AMINO  ACIDS    103 

As  a  general  rule  substances  containing  the  aromatic 
or  benzene  nucleus  do  not  readily  undergo  complete 
oxidation  in  the  organism,  the  benzene  nucleus  remain- 
ing unchanged.  The  amino  acids  derived  from  protein 
hydrolysis,  and  containing  this  nucleus,  namely,  tyro- 
sine, phenylalanine  and  tryptophane  do  suffer  complete 
disintegration,  the  benzene  nucleus  being  disrupted. 
There  are  at  least  two  ways  in  which  the  aromatic 
amino  acids  may  be  destroyed.  In  the  first  place  the 
following  series  of  reactions  may  occur — phenylalanine 
being  employed  as  a  specific  example.  Phenylalanine 
by  oxidative  deamination  is  first  changed  to  phenyl- 
pyruvic  acid : 


COOH 

a  ketone  acid 

COOH 

CH.NH, 

C  =  O 

CH, 

CH, 

C 

/\ 
HC        CH 

A 

HC        CH 

HC        CH 

\/ 
CH 

HC        CH 

\/ 
CH 

Phenylalanine 

P 

'henyl-pynivic  a( 

104 


THE  AMINO  ACIDS 


If  we  assume  that  the  next  step  is  the  simple  spHtting 
open  of  the  benzene  nucleus  of  phenyl-pyruvic  acid  its 
formula  may  be  written  as  follows : 


COOH 

COOH 

COOH 

C  = 

0 

CH, 

CH, 

CH, 

C= 

c  =  o 

C= 

by  cleavag-e 

of  CO3  this 

becomes 

CH 

CH, 

CH 
CH 

CH 

CH 

CH 

# 

CH 

CH 

C 

C  = 

0 

H 

H 

Phenyl-pyruvic 
acid  written  in 

Diacetic  acid 

open  chain  form 

By  the  splitting  in  two  of  this  chain  and  oxidation 
aceto-acetic  or  diacetic  acid  is  produced  which  in  turn 
may  be  directly  oxidized  to  CO2  and  HgO.  Secondly, 
employing  tyrosine  as  a  specific  example  we  may  follow 
it  through  the  following  changes : 


FURTHER  FATE  OF  AMINO  ACIDS    105 


OH 


OH 


by  oxidative 
deamination 


CH, 

CHNH, 

I 
COOH 


/ 


oxidation 

and 
rearrange- 
ment 


(Tyrosine) 

p.oxyphenyl 

amino-propionic 

acid 


(Ketone  acid) 

p.oxyphenyl 

pyroracemic 

acid 


HO 

/\ 

by 
CO. 

cleavage 

\ 
/ 

CH, 
C  =  O 
COOH 

\ 
/ 

H 

pyr 

ydroquino 
oracemic  j 

ne 
icid 

106  THE  AMINO  ACIDS 


HO 


OH  \ 


Acetone 
Bodies 


'^         CO,  +  H,0 
CH, 

COOH 

Homogentisic  acid 

In  these  transformations  homogentisic  acid  is  an 
important  intermediary  product  which  at  times  appears 
in  the  urine.  (See  Alkaptonuria.)  In  harmony  with 
the  idea  that  homogentisic  acid  is  an  intermediary 
product  in  the  decomposition  of  tyrosine  and  pheny- 
lalanine is  the  demonstration  that  the  liver  may  form 
acetone  from  homogentisic  acid. 

Little  definite  is  known  concerning  intermediary 
stages  in  the  fate  of  trytophane  in  the  human  body. 

Arginine  undoubtedly  undergoes  hydrolysis  by  the 
enzyme  arginase  yielding  urea  and  ornithine  and  the 
latter  may  also  yield  urea. 

NH, 

C  =  NH 

NH.CH,.CH,.CH,.CH.NH,.COOH  +  H^O  = 

Arginine 


FURTHER  FATE  OF  AMINO  ACIDS    107 


CH,.CH,.CH,.CH.COOH 

NH, 

C  =  O  +  NH,                  NH, 

NH, 

Omithine 

Synthesis  of  Amino  Acids 

It  has  been  demonstrated  that  alanine,  phenylalanine 
and  tyrosine  may  be  synthesized  in  the  liver  by  per- 
fusion of  ammonium  salts  of  ketone  acids. 

R.CH,.CO.COONH, 

Ammoniuin  salt  of  ketonic  acid 

OH 


R.CH,.Cf COOH         R.CH,.CH.NH,.COOH 


NH 

Imino  acid  hydrate  Amino  acid 

The  possibility  of  synthesis  of  amino  acids  in  this 
manner  renders  the  interpretation  of  metabolic  changes 
in  the  tissues  more  complex  than  ever  and  confers  upon 
the  organism  a  range  of  synthetic  powers  practically 
unlimited. 

Under  ordinary  circumstances,  however,  it  is  un- 
likely that  amino  acids  are  synthesized  to  a  great  extent 


108  THE  AMINO  ACIDS 

in  this  way.  The  question  has  been  tested  experiment- 
ally in  an  indirect  manner.  If  protein  with  a  certain 
amino  acid  lacking  is  fed  to  animals  it  is  reasonable 
to  assume  that  if  nitrogenous  equilibrium  can  be  main- 
tained a  synthesis  of  the  missing  amino  acid  must  have 
occurred.  From  experiments  planned  to  test  this 
hypothesis  it  has  been  shown  that  no  amino  acids  with 
the  exception  of  glycocoll  are  ordinarily  formed  by 
synthesis.  For  glycocoll  the  evidence  is  strongly  indic- 
ative of  synthesis.  As  a  rule  in  the  body  there  is 
about  5  per  cent  of  glycocoll  nitrogen  in  every  100 
grams  of  protein  nitrogen.  It  is  well  known  that 
benzoic  acid  ingested  is  united  with  glycocoll  to  form 
hippuric  acid — in  other  words,  benzoic  acid  feeding 
^-  robs  the  body  of  glycocoll.  If  benzoic  acid  is  fed  in 
sufficient  quantities  to  exhaust  the  possible  content  of 
glycocoll  preformed  in  the  tissues,  the  continued  forma- 
tion of  hippuric  acid  must  be  provided  for  by  glycocoll 
newly  formed  or  synthesized.  Hippuric  acid  does  con- 
tinue to  be  formed  under  these  circumstances  and 
hence  glycocoll  must  be  synthesized.  It  is  possible,  of 
course,  that  glycocoll  may  be  formed  from  the  trans- 
formation of  some  other  amino  acid,  as  by  cleavage  of 
a  long  change  amino  acid.  Another  evidence  in  favor 
of  the  synthesis  of  glycocoll  is  the  following — milk 
proteins  are  very  poor  in  glycocoll,  yet  suckling  ani- 
mals are  capable  in  a  short  time  of  building  up  in  their 
bodies  proteins  which  contain  far  more  of  this  amino 
'^^  acid  than  can  be  accounted  for  by  the  ingestion  of 
glycocoll  yielded  by  the  milk. 


FURTHER  FATE  OF  AMINO  ACIDS    109 

The  Relationship  Between  Carbohydrates  and 

Amino  Acids 

I.    The  Formation  of  Carbohydrate  from  Amino  Acids 

For  a  long  time  it  has  been  accepted  that  carbo- 
hydrate may  be  formed  from  ingested  protein.  To 
determine  the  mechanism  of  this  transformation  many 
experiments  have  been  carried  through  upon  animals. 
In  particular  the  formation  of  glycogen  from  ingested 
protein  has  been  subjected  to  experimentation  and 
although  the  consensus  of  opinion  would  indicate  that 
protein  may  give  rise  to  glycogen  formation,  the 
experimental  conditions  under  which  most  of  the  inves- 
tigations were  made  are  not  free  from  criticism.  The 
evidence  of  clinical  experience,  with  diabetes,  where 
fat  or  carbohydrate  ingestion  cannot  always  be  held 
responsible  for  the  large  amounts  of  sugar  passing 
through  the  kidneys  daily,  points  positively  to  protein 
as  the  source  of  the  carbohydrate  excreted.  In  agree- 
ment with  this  conception  is  the  observation  that  the 
urinary  nitrogen  and  sugar  excretion  in  the  patho- 
logical state  mentioned  run  along  parallel  lines. 

How  may  this  sugar  formation  be  explained?  One 
may  assume,  for  instance,  that  protein  contains  groups 
of  a  carbohydrate  nature  or  groups  closely  allied  to  the 
carbohydrates.  Although  it  must  be  accepted  that 
certain  proteins  do  contain  carbohydrate  groups,  the 
possession  of  such  groups  by  proteins  is  by  no  means 
universal,  and,  on  the  other  hand,  one  is  unwarranted 
in  stating  that  any  specific  protein  will  not  lead  to 


110  THE  AMINO  ACIDS 

sugar  production.  If  we  take  a  typical  protein  as  egg 
albumin,  and  then  on  the  assumption  that  all  the  nitro- 
gen present  is  eliminated  from  the  body  as  urea,  we 
find  left  over  a  large  carbon  residue,  the  so-called 
"carbon  moiety,"  of  the  protein  which  may  be  regarded 
as  material  capable  of  being  transformed  into  carbo- 
hydrate. This  potential  of  carbohydrate  forming 
material  must  gain  access  to  the  blood  stream,  hence 
to  the  tissues,  in  the  form  of  amino  acids,  in  accord- 
ance with  the  present-day  view  of  the  processes  of 
metabolism.  The  problem  under  consideration,  there- 
fore, resolves  itself  in  the  question,  "Are  amino  acids 
capable  of  being  transformed  into  carbohydrates?" 
The  most  convincing  work  in  this  direction  is  that  of 
Lusk  and  his  pupils.  They  have  administered  to  dogs 
rendered  diabetic  with  phlorhizin  various  amino  acids 
and  have  observed  that  some  yield  sugar  whereas 
others  fail  to  do  so.  In  their  experiments  the  relation- 
ship between  the  dextrose  and  nitrogen  of  the  urine, 
the  D :  N  ratio,  was  determined.  Under  suitable  con- 
ditions this  becomes  a  constant.  The  ingestion  of 
sugar  forming  substances  changes  this  constant  and 
any  change  serves  as  an  index  to  the  quantity  of  sugar 
formed  from  a  given  amount  of  substance  introduced. 
It  was  found  that  the  N-f ree  parts  of  glycocoll,  alanine, 
aspartic  acid,  and  glutamic  acid,  containing  respectively 
two,  three,  four,  and  five  carbon  atoms  may  be  either 
completely  or  partially  transformed  to  dextrose.  All 
of  the  glycocoll  and  all  of  the  alanine  were  converted 


FURTHER  FATE  OF  AMINO  ACIDS    111 

into  glucose,  whereas  three  of  the  carbon  atoms  con- 
tained in  aspartic  and  glutamic  acids  were  so  changed. 
The  stages  through  which  these  amino  acids  are 
carried  have  been  outlined  by  Lusk.  In  the  first  place 
it  is  assumed  that  the  initial  change  is  a  hydrolytic 
deamination  whereby  ammonia  is  formed  and  a 
hydroxy  group  is  added  to  the  denitrogenized  amino 
acid.  According  to  this  view  glycocoll  would  first  be 
changed  to  glycolic  acid,  which  on  reduction  would 
yield  glycolic  aldehyde,  three  molecules  of  which  will 
form  one  molecule  of  dextrose.  The  chemical  rela- 
tionships are  shown  below : 

CH,.NH,  CH^.OH 

3        I  +  H,0  =  3      I  -  3  O  = 

COOH  COOH 

Glycocoll  Glycolic  acid 


CH.GH 
5       I 
CHO 

Glycolic 
aldehyde 


CeHaOs 

Dextrose 


For  alanine  the  changes  undergone  are  a  direct  trans- 
formation into  lactic  acid  which  is  well  known  to  give 
rise  to  dextrose  production. 

CHs  CHg 

2     CHNH,  +  H,0     =     2     CHOH     =     C.HeOe 


COOH 

Alanine 


COOH 
Lactic  acid 


Dextrose 


112 


THE  AMINO  ACIDS 


With  aspartic  acid  only  three  of  the  carbon  atoms 
are  available  for  dextrose  formation,  the  remaining 
carbon  atom  being  changed  to  carbon  dioxide  in  ac- 
cordance with  the  following  reactions : 


COOH 

I 
CH, 

CHNH, 


+  H,0 


COOH 
Aspartic  acid 


COOH 

I 
CH, 

CH.OH 

2  CO, 
/3  lactic  acid 


C.H.O. 


Dextrose 


As  with  aspartic  acid  so  also  with  glutamic  acid  only- 
three  of  the  carbon  atoms  are  changed  to  dextrose,  the 
two  remaining  carbon  atoms  being  liberated  in  the 
form  of  acetic  acid. 


COOH 

CH, 

2       I 
CH, 

CHNH, 

COOH 

Glutamic  acid 


+  4  H,0 


COOH 
CH3 


CH.OH 
2  CHOH 

COOH 

Acetic  acid  and 
Glyceric  acid 


=     C.H«0, 


Dextrose 


Dakin  has  demonstrated  that  serine,  proline,  orni- 
thine and  arginine  are  all  capable  of  yielding  large 
amounts   of   sugar   when  given   to   glycosuric   dogs. 


FURTHER  FATE  OF  AMINO  ACIDS    113 

Apparently  arginine  is  the  only  amino  acid  with  more 
than  five  carbon  atoms  which  furnishes  glucose  freely. 
In  this  case  it  is  probable  that  the  sugar  comes  from 
the  ornithine  moiety  with  five  carbon  atoms,  into  which 
it  may  be  converted  by  the  action  of  arginase.  Lysine 
is  the  only  straight  chain  amino  acid  derivative  of 
protein  which  fails  to  yield  sugar.  Although  the  rela- 
tionship of  the  remaining  amino  acids  to  carbohydrate 
metabolism  is  less  definitely  established,  Lusk  has 
made  the  interesting  calculation  that  in  diabetes  sugar 
may  arise  from  protein  to  the  extent  of  nearly  60  per 
cent.  He  says,  "It  becomes  evident  that  there  may 
be  a  condition  of  nutrition  in  which  protein  is  used 
neither  for  repair  nor  for  growth,  but  simply  to  be 
diaminized  and  subsequently  to  act  like  fat  or  carbo- 
hydrate as  nutritive  materials  for  the  organism." 

2.     The  Formation  of  Amino  Acids  from  Carbo- 
hydrates 

The  formation  of  amino  acids  from  carbohydrate 
material  is  a  reaction  less  well  known  than  the  reverse 
process.  The  close  relationship  existing  between  lactic 
acid  and  carbohydrates  on  the  one  hand  and  lactic  acid 
and  alanine  on  the  other  suggests  the  ready  trans- 
formation of  glycogen  to  alanine  presumably  with 
lactic  acid  and  ammonium  pyruvate  as  intermediary 
products.  With  this  suggestion  in  mind  Embden  per- 
fused a  liver  rich  in  glycogen  and  found  that  alanine 
was  formed.    When,  however,  perfusion  of  a  glycogen- 


114  THE  AMINO  ACIDS 

free  liver  was  carried  through  alanine  was  not  present 
in  significant  quantities.  From  experiments  of  this 
nature  it  is  evident  that  the  metabolic  processes  con- 
cerned in  protein  metabolism  are  intimately  associated 
with  those  of  the  intermediary  metabolism  of  carbo- 
hydrate, and  further  that  at  times  at  least  protein  may 
serve  for  both  sources  of  nitrogen  and  carbonaceous 
material.  Protein,  therefore,  should  not  be  regarded 
in  the  strict  sense  merely  as  a  purveyor  of  the  nitrogen 
which  is  essential  for  life  processes.  It  is  much  more 
than  that,  as  has  been  demonstrated. 

Anomalies  of  Amino  Acid  Metabolism 
Alkaptonuria  • 

In  previous  pages  it  has  been  pointed  out  that  the 
normal  organism  is  capable  of  demolishing  the  benzene 
ring  as  found  in  tyrosine  and  phenylalanine  for  under 
normal  conditions  no  evidence  of  these  substances  can 
be  found  in  the  urine.  For  the  successive  steps  as- 
sumed to  occur  in  this  destruction  see  p.  105.  How- 
ever, there  are  certain  individuals  who  apparently  are 
unable  to  break  open  the  aromatic  ring  and  in  the  urine 
is  found  an  intermediary  decomposition  product,  homo- 
gentisic  acid.  Urine  containing  homogentisic  acid 
exhibits  a  tendency  to  turn  dark  on  exposure  to  the  air 
and  may  show  a  strong  reducing  action.  This  condi- 
tion, known  as  alkaptonuria,  is  of  rare  occurrence  and 
may  last  through  life  without  affecting  the  health  of 
the  individual.     It  can  scarcely  be  regarded  as  of 


FURTHER  FATE  OF  AMINO  ACIDS    115 

pathological  nature,  but  should  be  looked  upon  rather 
as  an  anomaly  of  metabolism  and  is  generally  con- 
sidered as  being  hereditary  in  origin.  It  occurs  oftener 
in  man  than  in  woman  and  blood  relationship,  as  first 
cousins,  predisposes  to  the  condition. 

Whenever  homogentisic  acid  is  present  in  the  urine, 
it  is  there  in  relatively  large  amounts  for  the  anomaly 
is  an  absolute  one,  that  is,  apparently  homogentisic 
acid  formed  is  destroyed  by  the  normal  organism. 
The  relationship  of  tyrosine  phenylalanine  and  homo- 
gentisic acid  are  shown  below : 


OH 


HO 


OH 


\X 

CH, 

CH, 

CH, 

COOH 

CHNH, 

CH.NH, 

COOH 

COOH 

Pyrosine 

Homogentisic  acid 

Phenylalanine 

The  significance  of  alkaptonuria  in  connection  with 
the  metabolism  of  the  amino  acids  is  that  the  appear- 
ance of  homogentisic  in  the  urine  of  alkaptonurics 
gave  the  first  hint  as  to  the  probable  transformations  ^ 
occurring  in  the  demolition  of  the  benzene  radical 
found  in  tyrosine  and  phenylalanine.    That  homogen- 


116  THE  AMINO  ACIDS 

tisic  acid  is  a  step  in  the  degradations  of  these  amino 
acids  has  been  doubted  by  Dakin  who  beUeves  that 
there  is  in  these  subjects  an  abnormal  formation  of 
homogentisic  acid  as  well  as  an  inability  to  destroy  it 
once  formed.  In  accord  with  this  idea  he  has  shown 
how  tyrosine  and  phenylalanine  may  be  destroyed  with- 
out homogentisic  acid  as  an  intermediary  product  (see 
p.  103).  On  the  other  hand,  it  has  been  demonstrated 
by  Abderhalden  that  normal  individuals  may  eliminate 
homogentisic  acid  when  excessive  quantities  of  tyrosine 
are  fed.  Also  the  administration  of  tyrosine  or  phenyl- 
alanine, or  of  foods  rich  in  tyrosine,  that  is  proteins, 
causes  a  significant  increase  in  the  excretion  of  homo- 
gentisic acid.  If  alkaptonurics  live  on  a  protein-free 
diet  for  short  periods  of  time  the  excretion  of  homo- 
gentisic acid  is  markedly  diminished,  but  does  not  dis- 
appear entirely.  Undoubtedly  the  aromatic  amino 
acids  formed  from  tissue  metabolism  do  not  suffer 
destruction  to  any  greater  extent  than  those  introduced 
into  the  blood  from  the  food  protein.  Why  an  alkap- 
tonuric  individual  fails  to  destroy  the  tyrosine  radicle 
is  still  a  matter  of  conjecture. 

Cystinuria  and  Diaminuria 

,  Under  ordinary  circumstances  cystine,  the  sulphur 
bearing  amino  acid  fails  to  appear  in  the  excreta,  prob- 
ably undergoing  extensive  destruction.  The  sulphur 
is  oxidized  to  sulphuric  acid  and  eliminated  as  a  sul- 
phate in  the  urine.  In  certain  individuals,  however, 
cystine  appears  in  the  urine  and  owing  to  its  relatively 


FURTHER  FATE  OF  AMINO  ACIDS    117 

slight  solubility  is  deposited  as  hexagonal  crystals.  It 
may  also  form  cystine  concretions  in  the  bladder.  The 
condition  of  cystinuria  with  that  of  alkaptonuria  must 
be  regarded  as  an  anomaly  of  metabolism.  Cystinuria 
appears  to  be  a  distinctly  hereditary  condition  since 
it  may  appear  in  families  for  many  generations,  and 
apparently  follows  the  Mendelian  law  of  heredity. 
Like  alkaptonuria  it  is  found  oftener  in  males  than  in 
females ;  it  seems  to  lead  to  no  pathological  symptoms 
other  than  the  formation  of  concretions.  There  is 
probably  no  complete  failure  to  destroy  cystine  since 
only  a  portion  of  the  cystine  from  protein  ingested 
reappears  in  the  urine.  Undoubtedly  a  part  of  the 
cystine  is  catabolized  in  a  normal  manner.  From  the 
fact  that  cystinuria  persists  in  the  absence  of  protein 
intake  and  further  that  cystine  fed  to  cystinurics  fails 
to  appear  in  the  urine,  the  conclusion  may  be  reached 
that  the  urinary  cystine  has  its  origin  in  that  formed 
during  catabolism  of  the  tissues.  It  may  be  possible 
that  in  these  subjects  there  is  only  a  limited  capacity 
for  destroying  amino  acids  in  general  for,  in  addition 
to  cystine,  leucine  and  tyrosine  have  been  found  in 
some  cases. 

At  times  the  diamines  cadaverine  and  putrescine 
formed  by  putrefaction  in  the  intestine  may  also  be 
present  in  the  urine  of  cystinurics.  The  diamines  are 
significant  in  that  they  are  two  of  the  so-called 
ptomaines.  Putrescine  and  cadaverine  occur  also  in 
diseased  conditions  of  the  intestinal  tract,  thus  they 
may  be  foimd  in  various  infections,  in  cholera,  dysen- 


118  THE  AMINO  ACIDS 

tery,  gastro-enteritis,  etc.  Their  origin  in  putrefaction 
of  protein  decomposition  products  together  with  their 
appearance  in  the  urine  of  cystinurics  led  to  the  view 
that  the  cystine  in  the  cases  mentioned  had  a  Hke  source. 
This,  however,  has  been  shown  to  be  incorrect.  A 
second  view  was  that  diamines  interfered  with  sulphur 
oxidation  in  the  organism,  hence  the  appearance  of  the 
unoxidized  cystine.  This  idea  has  been  shown  to  be 
untrue  for  cystinuria  may  occur  in  the  absence  of  the 
diamines,  and  the  administration  of  diamines  has  no 
influence  upon  the  output  of  cystine  in  the  urine.  "In 
intestinal  disturbances,  it  is  probable  that  these  com- 
pounds are  the  result  of  bacterial  activity — indeed,  they 
may  be  the  metabolic  end-products  eliminated  by  bac- 
teria. In  cystinuria,  however,  it  is  possible  that  a  dif- 
ferent explanation  for  diaminuria  is  pertinent.  It  may 
be  assumed,  for  instance,  that  in  the  beginning  cysti- 
nuria and  diaminuria  are  brought  about  through  a 
similar,  or  indeed  the  same  cause,  or  causes,  for 
example,  a  gradually  changing  type  of  metabolism 
induced  by  some  unknown  agency,  resulting  in  an 
anomaly  of  metabolism.  If  the  anomaly  is  slight  in 
character,  cystine  alone  is  eliminated  as  a  result, 
whereas  if  the  change  in  metabolism  is  sufficiently  pro- 
nounced diamines  are  also  excreted.  If  this  assump- 
tion is  accepted  it  is  easy  to  explain  why  in  some  cases 
of  cystinuria  the  diamines  are  absent,  and  that  gradu- 
ally one  or  both  of  these  compounds  disappear,  that 
cystinuria  persists,  but  that  cystinuria  does  not  cease 
and  leave  diaminuria." 


FURTHER  FATE  OF  AMINO  ACIDS    119 

References  to  Literature 

Fate  of  Amino  Acids 

Dakin:  Oxidations  and  Reductions  in  the  Animal  Body.    1912. 
Dakin:  Journal  of  Biological  Chemistry.    1913,  14,  p.  321. 
Lusk:  Journal  of  the  American  Chemical  Society.     1910,  32, 

p.  671. 
Lusk:  Ergebnisse  des  Physiologic.    1912,  12,  p.  315.     [Phlor- 

hizin  glycosuria.] 

Alkaptonuria 

Neuherg:  Oppenheimer's  Hanbuch  der  Biochemie.    1910,  IV, 

2,  p.  353. 
Wells:  Chemical  Pathology.  1914. 

Cystinuria  and  Diaminuria 

Neuherg:  Oppenheimer's  Handbuch  der  Biochemie.    1910,  IV, 

2,  p.  338. 
Underhill:  Middleton  Goldsmith  Lecture  for  1911.    Archives 

of  Internal  Medicine.    1911,  8,  p.  7  and  p.  17. 
Wells:  Chemical  Pathology.    1914. 


CHAPTER  VII 

THE  AMINO  ACIDS  IN  RELATION  TO  THE 
SPECIFIC  DYNAMIC  ACTION  OF 
PROTEINS 

The  view  has  been  held  for  many  years  that  the 
ingestion  of  protein  increases  the  power  of  the  body 
cells  to  metabolize  materials  brought  to  them.  More 
recently  it  has  been  shown  by  Rubner  that  under 
suitable  conditions  each  foodstuff  is  capable  of  exert- 
ing a  specific  accelerating  influence  upon  the  energy 
metabolism.  In  order  to  maintain  life  Rubner  believes 
that  a  fixed  requirement  of  energy  is  necessary.  When 
the  organism  is  fasting,  the  essential  energy  require- 
ment may  be  furnished  from  the  tissues  of  the  organ- 
ism itself.  After  their  ingestion  the  foodstuffs  are 
changed  in  various  ways  with  the  evolution  of  heat 
until  finally  they  are  transformed  into  materials  which 
are  capable  of  supporting  vital  phenomena.  The  final 
products  formed  may  be  employed  for  the  replacement 
of  the  substances  oxidized  during  fasting.  The  heat 
produced  in  the  formation  of  these  compounds  is 
added  to  the  heat  produced  for  the  maintenance  of 
vital  processes,  and  the  total  heat  production,  there- 
fore, exceeds  that  found  in  starvation.    This  increased 


DYNAMIC  ACTION  OF  PROTEINS      121 

heat  production  induced  by  each  foodstuff  is  different, 
and  is  specific  for  each  foodstuff.  Rubner,  therefore, 
has  named  this  effect  "specific  dynamic  action." 

It  has  been  shown  under  the  correct  conditions  that  if 
the  energy  metabohsm  of  a  fasting  dog  be  represented 
as  100  calories,  food  must  be  given  in  the  following 
amounts  to  prevent  body  loss,  106  calories  of  sugar, 
or  114  calories  of  fat,  or  140  calories  of  protein.  In 
an  experiment  by  Rubner  it  was  found  that  a  fasting 
man  metabolized  2042  calories.  After  the  ingestion 
of  2450  calories  of  sugar  he  metabolized  2087.  When 
given  2450  calories  of  meat  2566  calories  were  metab- 
olism. Whatever  the  cause  of  the  greater  metabolism 
of  protein  ingestion  it  is  believed  that  there  is  produced 
a  liberation  of  energy  which  cannot  be  used  by  the 
tissues  in  support  of  their  activities  but  it  is  possible 
that  it  may  contribute  to  the  maintenance  of  body 
temperature.  On  a  mixed  diet  this  liberation  of  heat 
unavailable  for  energy  purposes  is  not  of  great  sig- 
nificance in  the  total  metabolism  since  it  increases  the 
metabolism  of  energy  less  than  one  tenth  on  a  mainte- 
nance diet  above  that  when  no  food  is  eaten. 

Another  explanation  for  the  increased  energy  metab- 
olism after  the  ingestion  of  foodstuffs  has  been  put 
forward  by  Zuntz,  who  ascribes  this  effect  to  the 
mechanical  work  of  the  intestinal  canal  (Darmarbeit) 
performed  during  digestion  and  assimilation.  Such  a 
theory  would  seem  to  fit  in  well  with  the  greater 
specific  dynamic  action  of  protein  and  its  probably 
greater   difficulty   of    digestion   in   comparison   with 


122  THE  AMINO  ACIDS 

sugar.  To  test  the  Zuntz  hypothesis  Benedict  has 
attempted  to  produce  the  effects  of  food  ingestion 
such  as  augmented  mechanical  work  through  increased 
peristalsis  produced  by  large  doses  of  purgatives,  as 
sodium  sulphate.  In  spite  of  greatly  increased  peris- 
talsis the  administration  of  sodium  sulphate  failed  to 
show  any  measurable  increase  in  metabolism.  "In  the 
belief  that  when  the  intestine  is  full  of  partly  digested 
food  products  and  epithelial  debris,  the  amount  of 
mechanical  work  thereby  incurred  might  be  greater 
than  that  involved  in  several  powerful  peristaltic 
waves,  experiments  were  made  in  which  relatively 
large  amounts  of  agar-agar  were  ingested,  thus  pro- 
ducing a  bulky,  voluminous  stool.  The  agar-agar 
being  practically  non-oxidizable,  there  was  no  great 
complication  due  to  the  combustion  of  carbohydrate 
from  the  agar-agar.  With  the  agar-agar  it  is  reason- 
able to  assume  that  there  must  have  been  an  extensive 
segmentation  process  as  well  as  peristaltic  waves. 
But  even  under  these  conditions  on  the  ideally  con- 
trolled experiments  there  was  absolutely  no  increase 
in  metabolism.  In  so  far,  then,  as  the  experiments  on 
men  show  with  controlled  conditions,  the  work  of 
peristalsis  and  probably  of  segmentation  is  not  suffi- 
cient to  be  measured  in  the  great  daily  energy  trans- 
formation of  the  body.  It  is  impossible  to  think  of 
muscular  activity  of  any  kind  taking  place  without 
some  slight  increased  metabolism,  but  the  amount 
involved  in  intestinal  activity  must  be  so  small  as  to 


DYNAMIC  ACTION  OF  PROTEINS      123 

be  entirely  negligible  in  the  extensive  energy  trans- 
formations for  body  maintenance." 

These  results  are  in  accord  with  conclusions  reached 
earlier  by  Rubner  who  denied  that  the  work  of  diges- 
tion and  assimilation  could  be  held  responsible  for  the 
effects  observed  after  food  ingestion. 

Benedict  is  of  the  opinion  that  from  the  food  sub- 
stances are  absorbed  which,  carried  by  the  blood,  stimu- 
late cells  to  greater  activity,  and  he  further  indicates 
that  these  unknown  bodies  are  of  an  acid  character. 
From  his  extensive  investigation  on  the  influence  of 
amino  acids  upon  metabolism,  that  is,  their  specific 
dynamic  action,  Lusk  believes  that  the  explanation  of 
Rubner  as  to  the  cause  of  specific  dynamic  action  must 
be  revised.  "Amino  acids  act  as  stimuli  upon  the 
cells,  raising  their  power  to  metabolism.  They  may 
act  instead  of  nerve  stimuli  when  increased  heat  pro- 
duction is  required  in  the  presence  of  external  cold — 
the  chemical  regulation  of  temperature  of  Rubner. 
The  energy  liberated  in  response  to  these  stimuli  may 
be  supplied  by  carbohydrate  or  fat.  When  fat  and 
carbohydrate  are  given  separately  or  together  there 
may  be  an  increased  heat  production  on  account  of 
the  increase  in  the  quantity  of  materials  available  for 
the  nutrition  of  the  cells.  With  the  cessation  of 
absorption  and  the  return  of  the  blood  to  the  compo- 
sition it  possessed  before  food  was  taken  the  metabo- 
lism falls  to  its  basal  value."  When  glucose  and  an 
amino  acid,  as  alanine,  are  given  together  the  metab- 
olism is  increased  to  a  point  where  the  resultant  effect 


124  THE  AMINO  ACIDS 

is  nearly  equal  to  the  sum  of  the  two  individual  influ- 
ences. This  indicates  a  distinct  difference  between  the 
cause  of  the  specific  dynamic  action  of  glucose  and 
that  of  alanine.  Two  types  of  processes  are  here  sug- 
gested, namely,  a  metabolism  of  plethora  and  amino 
acid  stimulation.  Carbohydrate  or  fat  metabolites 
which  are  being  absorbed  from  the  intestine  into  the 
blood  bring  about  a  metabolism  of  plethora.  In  the 
metabolism  of  plethora  the  influx  of  carbohydrate  or 
fat  enables  the  cells  to  oxidize  at  a  higher  level  through 
the  increased  mass  action  of  food  particles  which  are 
available.  (Lusk.)  A  recent  attempt  by  Lusk  to  ex- 
plain "amino  acid  stimulation"  of  the  cells  has  resulted 
in  the  conclusion  that  some  at  least  of  the  amino  acids 
even  when  they  are  not  oxidized  "yield  products  of 
metabolism,  either  hydroxy  or  ketone  acids  which  act 
as  stimuli  to  induce  higher  oxidation  in  the  organism. 
This  is  the  conclusive  proof  of  a  true  chemical  stimu- 
lation of  protoplasm  within  the  mammalian  organism. 
It  explains  the  specific  dynamic  action  of  protein." 

As  has  been  shown  repeatedly  throughout  this  book 
the  effects  characteristically  produced  by  protein  are 
gradually  being  ascribed  as  a  function  of  the  amino 
acids.  The  amino  acids  therefore  may  be  regarded 
not  alone  as  pabulum  for  the  restoration  of  depleted 
cells  but  must  also  be  looked  upon  as  playing  a  distinct 
and  significant  role  in  the  rate  or  extent  of  cellular 
metabolism. 


DYNAMIC  ACTION  OF  PROTEINS      125 

References  to  Literature 

Lusk:  The  Science  of  Nutrition.    Second  Ed.    1909. 

Lusk:  Journal  of  Biological  Chemistry.    1915,  20,  p.  555. 

Rubner:  Zuntz,  Benedict,  Lusk.  Trans,  of  the  15th  Inter- 
national Congress  on  Hygiene  and  Demography.  1913, 
Part  2. 

Sherman:  Chemistry  of  Food  and  Nutrition.    1911. 


CHAPTER  VIII 

THE  AMINO  ACIDS  AND  SIMPLER  NITRO- 
GENOUS COMPOUNDS  AS  FOODSTUFFS 

Value  of  Amino  Acids  as  Foodstuffs 

The  dictum  of  Liebig  that  the  animal  organism  is 
endowed  with  very  limited  capacities  for  processes  of 
synthesis  was  accepted  for  a  great  many  years  with- 
out serious  question.  A  partial  reason  for  this  assump- 
tion is  to  be  found  in  the  great  difficulties  to  be  over- 
come in  putting  the  question  to  experimental  proof. 
The  discovery  of  erepsin  by  Cohnheim  with  the  con- 
sequent readjustment  of  ideas  relative  to  the  extent 
and  character  of  digestion  processes  and  the  form  of 
products  absorbed  casts  doubt  upon  the  inability  of  the 
animal  body  to  synthesize.  It  was  reasonable  to 
assume  that  the  disintegration  of  the  protein  molecule 
to  the  stage  of  amino  acids  is  with  a  purposeful  object 
and  that  the  absorption  of  such  relatively  small  com- 
pounds as  amino  acids  predicates  the  probability  that 
synthesis  must  occur  if  the  organism  is  obliged  to 
reconstruct  new  protoplasm  to  replace  that  worn  away 
through  the  many  metabolic  activities. 

About  a  dozen  years  ago  the  first  attempt  to  deter- 
mine the  possibility  of  positive  protein  synthesis  was 


NITROGENOUS  COMPOUNDS  127 

made  by  Loewi.  This  investigator  allowed  protein  to 
digest  until  it  no  longer  gave  a  reaction  with  the  biuret 
test  an  indication  that  products  possessed  of  a  protein 
nature  had  all  been  reduced  to  a  lower  stage.  Upon 
feeding  this  mixture  of  amino  acids  together  with 
fat  and  carbohydrate  to  a  dog,  Loewi  demonstrated 
that  nitrogen  in  the  form  contained  in  his  digestion 
mixture  was  not  alone  capable  of  maintaining  the  life 
of  the  animal,  but  furthermore  kept  it  in  a  state  of 
nitrogenous  equilibrium,  a  retention  of  nitrogen  to- 
gether with  an  increase  in  weight  being  observed. 
The  experiments  of  Loewi  were  quickly  followed  by 
those  of  Henderson  and  Dean,  who  were  the  first  to 
employ  digestion  products  formed  through  the  action 
of  acids  rather  than  by  ferments.  They  found  nitro- 
gen retention  but  were  uncertain  whether  it  signified 
protein  synthesis.  As  a  result  of  many  succeeding 
investigations  it  soon  became  clear  that  the  power  of 
the  digestion  products  to  replace  body  tissue  depended 
upon  the  manner  in  which  such  products  were  formed. 
To  put  it  differently,  digestion  products  formed  from 
protein  by  the  agency  of  enzymes  were  fully  capable 
of  supplying  to  the  body  its  necessary  quota  of  nitro- 
gen whereas  those  products  obtained  from  proteins 
through  the  action  of  acids  could  not  take  over  so 
completely  this  function  but  were  regarded  as  of 
value  inasmuch  as  they  could  be  looked  upon  as  being 
"protein  sparers."  In  this  connection  it  may  be  well 
to  cite  some  experiments  of  Abderhalden  and  Rona 
with  mice.    To  these  mice  were  fed  different  prepara- 


128  THE  AMINO  ACIDS 

tions  of  casein  together  with  sugar.  To  one  series  of 
mice  unchanged  casein  was  fed,  to  a  second,  casein 
that  had  been  digested  with  pancreatin  for  a  period 
of  two  months,  a  third  series  received  casein  digested 
for  one  month  with  pepsin-hydrochloric  acid  and  then 
for  two  months  with  pancreatin,  the  fourth  series 
were  fed  with  casein  hydrolyzed  for  ten  hours  with 
25  per  cent  sulphuric  acid.  The  results  showed  that 
those  animals  fed  with  casein  digested  with  pancreatin 
for  two  months  and  those  given  unchanged  casein 
lived  about  the  same  length  of  time.  Mice  fed  on  the 
other  two  preparations  lived  shorter  periods  of  time. 
Other  investigators  obtained  similar  results.  An 
interesting  controversy  now  arose  as  to  the  reason 
for  the  specific  difference  between  products  formed 
by  enzymes  and  those  resulting  from  acid  hydrolysis. 
Abderhalden  and  Rona  from  their  work  cited  above 
put  forth  the  hypothesis  that  the  difference  in  the 
two  products  lay  in  their  content  of  polypeptides. 
According  to  this  view  digestion  by  ferments  results 
in  the  presence  of  considerable  amounts  of  fairly 
complex  polypeptides  which  serve  as  nuclei  for  the 
synthesis  of  new  protein  material.  Hydrolysis  by  acid, 
however,  carries  the  digestion  beyond  the  stage  of 
polypeptides,  hence,  no  nuclei  for  synthesis  are  present 
and  the  inability  of  acid  digestion  mixtures  to  fully 
serve  as  nitrogenous  pabulum  is  explained.  In  sup- 
port of  their  hypothesis  they  offer  the  observation  that 
the  casein  preparation  formed  by  pancreatin  action 
contained  only  16  per  cent  of  polypeptides,  that  of  the 


NITROGENOUS  COMPOUNDS  129 

pepsin-hydrochloric  acid  mixture  further  subjected 
to  the  influence  of  pancreatin  contained  only  half  as 
much  polypeptides,  whereas  from  that  formed  by  acid 
hydrolysis  polypeptides  were  entirely  absent.  Later, 
however,  it  was  shown  by  Abderhalden  and  his  co- 
workers that  the  varying  content  of  polypeptides  can- 
not be  the  sole  reason  for  the  differences  observed  in 
the  two  classes  of  products  in  their  ability  to  supply 
the  nitrogenous  needs  of  the  body,  for  a  dog  was  kept 
alive  for  thirty-eight  days  and  the  only  supply  of  nitro- 
gen was  in  a  digestion  mixture  containing  only  amino 
acids.  Again,  a  young  dog  gained  weight  and  retained 
nitrogen  in  completely  digested  meat  and  a  bitch  was 
kept  in  nitrogenous  equilibrium  during  lactation  with 
meat  digested  to  the  amino  acid  stage.  Abderhalden 
and  London  were  able  to  maintain  a  dog  with  an  Eck 
fistula  (the  liver  shunted  out  of  the  portal  circulation) 
on  fully  digested  meat.  From  this  experiment  they 
further  concluded  that  the  liver  could  play  a  small 
role  only  in  protein  synthesis  and  used  these  results 
as  support  for  their  view  that  protein  synthesis  occurs 
during  absorption. 

From  the  work  of  Henriques,  Abderhalden  and 
others  it  soon  became  evident  that  the  difference  in 
nutritive  value  between  ferment  and  acid  hydrolysis 
products  could  not  be  ascribed  wholly  to  the  presence 
or  absence  of  polypeptides.  Upon  closer  investiga- 
tion it  developed  that  the  failure  of  acid  hydrolytic 
products  to  meet  nutritive  requirements  satisfactorily 
could  be  explained  by  the  fact  that  during  acid  hydroly- 


130  THE  AMINO  ACIDS 

sis  tryptophane,  unquestionably  one  of  the  most  impor^ 
tant  of  the  amino  acids,  is  destroyed.  The  proof  that 
herein  lies  the  true  explanation  was  furnished  by 
Abderhalden  and  Frank,  who  succeeded  in  maintaining 
dogs  in  nitrogenous  equilibrium  on  meat  completely 
hydrolyzed  by  acid  to  which  had  been  added  a  small 
amount  of  tryptophane. 

After  the  demonstration  that  the  nitrogenous  nutri- 
tive requirements  of  the  organism  can  be  supplied  by 
a  mixture  consisting  of  the  products  of  hydrolysis, 
whether  by  acids  or  enzymes,  an  attempt  was  made 
by  Abderhalden  to  support  a  dog  in  nitrogen  equilib- 
rium on  an  artificial  mixture  of  amino  acids  to  which 
were  added  carbohydrate  and  fat.  The  successful  out- 
come of  the  investigation  led  Abderhalden  to  declare 
that  the  animal  organism  is  capable  of  forming  all 
the  tissue  constituents  out  of  the  simplest  derivatives 
of  the  proteins.  Inasmuch  as  carbohydrate  and  fat 
may  be  prepared  synthetically,  as  may  some  of  the 
amino  acids,  the  problem  of  the  artificial  production 
of  foodstuffs  is  solved  according  to  Abderhalden, 
who  says  that  such  a  possibility  is  limited  only  by  the 
question  of  sufficient  funds.  From  these  observations 
it  becomes  evident  that  mixtures  of  amino  acids  are 
fully  capable  of  supplying  the  nitrogenous  needs  of 
the  organism  when  applied  to  the  lower  animals.  An 
opportunity  was  afforded  Abderhalden  and  his  co- 
workers to  extend  this  type  of  investigation  to  man. 
A  boy  with  a  stricture  of  the  oesophagus  on  whom 
gastrotomy  had  been  performed  was  the  subject.    To 


NITROGENOUS  COMPOUNDS  131 

him  was  given  per  rectum  a  mixture  of  protein  (meat) 
digestion  products  obtained  through  the  combined 
action  of  trypsin  and  erepsin.  The  experiment  was 
continued  for  fifteen  days  and  during  this  period 
nitrogen  equihbrium  was  maintained,  the  body  weight 
increased  and  the  general  condition  of  the  subject  was 
excellent. 

This  brief  review  of  the  salient  features  of  the 
problem  leads  to  but  one  conclusion,  namely,  that  the 
amino  acids  must  be  regarded  as  foodstuffs  capable 
of  supplying  the  nitrogenous  needs  of  the  organism, 
and  that  the  chief  factors  to  be  taken  into  account 
with  regard  to  the  nutritive  value  of  any  protein  or 
proteins  are  the  character  and  the  extent  of  the  amino 
acids  contained  therein. 


The  Value  of  Amides  and  Ammonium  Salts  as 

Foodstuffs 

The  nutritive  value  of  various  simple  nitrogenous 
compounds  has  been  a  subject  for  investigation  for 
many  years.  This  is  especially  true  for  such  sub- 
stances as  the  amides  and  has  been  of  particular  inter- 
est to  those  concerned  with  agricultural  problems 
since  in  the  food  of  herbivora  amides  may  play  an 
important  role.  From  the  viewpoint  of  nutrition  in 
general,  the  proof  that  animals  may  thrive  on  amides 
or  other  simple  nitrogenous  compounds  supplied  as 
sources  of  nitrogen  carries  with  it  indirect  evidence  of 


132  THE  AMINO  ACIDS 

the  transformation  of  these  substances  into  amino 
acids — in  other  words,  amino  acid  synthesis  occurs. 

Particular  attention  has  been  paid  to  the  deter- 
mination of  the  value  of  asparagine  as  a  source  of 
nitrogen,  one  of  the  first  investigators  being  Mer- 
cadente,  who  believed  that  protein  formation  could 
take  place  from  asparagine,  especially  in  plants. 
Sachse  believed  that  protein  was  formed  from  aspara- 
gine by  the  simple  addition  of  fatty  aldehydes.  On 
the  other  hand,  Loewi  thought  that  in  the  presence  of 
carbohydrates  protein  was  formed  from  asparagine 
by  a  series  of  condensations.  Zuntz  suggested  that  in 
herbivora  asparagine  was  built  up  into  protein  by 
bacteria  in  the  intestine  previous  to  utilization.  This 
latter  view  has  been  supported  by  numerous  investi- 
gators, some  of  whom  state  that  protein-forming  bac- 
teria are  widely  distributed  in  nature  and  may  act 
very  efficiently  and  quickly  when  in  suitable  envi- 
ronment. The  evidence  available  seems  to  speak 
strongly  in  favor  of  the  view  that  asparagine  may 
serve  as  source  of  nitrogen  or  act  at  least  as  a  protein 
sparer,  as  much  as  two-thirds  of  the  protein  in  the 
diet  of  herbivora  being  replaceable  by  asparagine.  On 
the  other  hand,  several  investigators  claim  that  aspara- 
gine cannot  take  the  place  of  protein  at  all,  hence, 
cannot  be  used  as  a  source  of  nitrogen  and  that  even 
the  degree  of  protein  sparing  action  that  may  be 
exhibited  by  this  amide  is  extremely  limited. 

In  a  comparable  manner  it  has  been  suggested  that 
various  ammonium  salts  may  also  replace  protein  to 


NITROGENOUS  COMPOUNDS  133 

a  certain  extent  at  least,  or  act  as  protein  sparers. 
The  problem  of  the  utilization  of  ammonium  salts 
subjected  to  much  experimentation  in  the  past  has 
recently  been  revived  through  the  work  of  Grafe  and 
Schlapfer  who  have  asserted  that  ammonium  salts, 
urea,  and  even  nitrates  may  serve  as  sources  of  nitro- 
gen for  the  animal  organism,  and  they  regard  the 
utilization  as  indicated  by  nitrogen  retention  as  a 
process  of  amino  acid  synthesis.  These  results  have 
been  assailed  by  others  on  the  ground  that  the  observed 
retention  of  nitrogen  as  a  result  of  feeding  the  above- 
mentioned  compounds  may  be  explained  in  other  ways 
than  as  a  proof  of  amino  acid  synthesis.  "There  are 
several  ways  in  which  they  may  be  assumed  to  behave 
in  the  organism.  In  the  first  and  foremost  instance 
they  may  serve  as  pabulum  for  the  alimentary  bac- 
teria, which  in  turn  are  destroyed  in  large  numbers 
in  the  digestive  tract  and  can  furnish  a  yield  of  per- 
fect protein  synthesized  from  simple  compounds  like 
urea  and  the  salts  of  ammonia.  It  is  generally  ad- 
mitted that  in  certain  species  like  the  herbivora,  in 
which  bacterial  processes  have  a  free  play  in  the 
gastro-intestinal  tube,  the  contribution  of  dead  bac- 
terial bodies  to  the  intake  is  by  no  means  negligible." 
"The  feeding  of  urea  or  ammonium  salts  might  lead 
to  an  apparent  nutritive  advantage  by  depressing  or 
inhibiting  the  usual  breaking  down  of  nitrogenous 
compounds  in  metabolism.  This  would  accord  with 
the  belief  that  the  products  of  cellular  waste  them- 
selves tend  to  impede  cellular  metabolism.     Now  that 


134  THE  AMINO  ACIDS 

the  synthesis  of  amino  acids  from  ammonia  and  car- 
bohydrates has  been  accompHshed  directly  or  in- 
directly in  the  laboratory,  the  possibility  of  a  similar 
reaction  in  the  body  must  be  reckoned  with.  Finally, 
the  alleged  utilization  of  urea  and  other  simple  nitro- 
gen derivatives  may  merely  be  an  instance  of  unsus- 
pected retention  and  delayed  excretion.  Even  so 
soluble  a  salt  as  an  iodide  may  not  be  entirely  recov- 
ered in  the  excreta  until  several  days  after  its  adminis- 
tration has  been  stopped.  Surely  no  one  would  look 
on  the  temporary  deficit  as  an  indication  of  nutritive 
'utilization'  of  the  foreign  salt." 

Various  possibilities  therefore  present  themselves  in 
the  interpretation  of  the  alleged  utilization  of  these 
simple  nitrogenous  substances.  The  influence  of 
alimentary  bacteria  may  be  eliminated  by  parenteral 
feeding  of  the  compounds,  which,  however,  has  not 
been  feasible  until  recently  when  Henriques  succeeded 
in  devising  a  method  whereby  a  slow  constant  stream 
of  nutritive  solution  may  be  intravenously  introduced 
into  the  body.  Subjecting  utilization  of  urea  and 
ammonium  salts  to  the  test  by  means  of  this  device, 
Henriques  and  Anderson  have  demonstrated  that  no 
permanent  retention  of  these  nitrogenous  compounds 
occurred.  It  is  therefore  exceedingly  improbable  that 
the  body  itself  is  in  a  position  to  transform  these  sub- 
stances into  amino  acids.  Amino  acid  synthesis  is 
not  an  easy  task  for  the  organism  nor  is  there  evidence 
that  even  the  transformation  of  one  amino  acid  to 
another  is  accomplished  to  any  extent.    The  organism 


NITROGENOUS  COMPOUNDS  135 

must  have  ready  formed  amino  acids  supplied  to  it 
in  sufficient  quantity  and  variety  if  it  is  to  accomplish 
its  task  of  tissue  building. 


References  to  Literature 

Amino  Acids  as  Foodstuffs 

Ahderhalden:  Zeitschrift  fiir  physiologische  Chemie.    1912,  77, 

p.  22. 
Cathcart:  The  Physiology  of  Protein  Metabolism.     1912. 
Henriques    and    Anderson:    Zeitschrift    fiir    physiologische 

Chemie.    1913,  88,  p.  357. 
Liithje:  Ergebnisse  des  Physiologie.    1908,  7,  p.  795. 
Cathcart:    The    Physiology    of    Protein    Metabolism.      1912. 

[Amides.] 

Ammonium  Salts 

Ahderhalden:   Zeitschrift   fiir   physiologische   Chemie.     1912, 
78,  p.  1,  and  1912,  82,  p.  1. 

Grafe  and  Schldpfer:  Zeitschrift  fur  physiologische  Chemie. 

1912,  77,  p.  1. 
Henriques    and    Anderson:     Zeitschrift    fiir    physiologische 

Chemie.      1914,  92,  p.  21. 
Pescheck:  Biochemische  Zeitschrift.    1912,  45,  p.  244. 
Underhill:  Journal  of  Biological  Chemistry.     1913,  15,  p.  327 

and  p.  337. 
Underhill  and  Goldschmidt:  Journal  of  Biological  Chemistry. 

1913, 15,  p.  341. 


CHAPTER  IX 

THE  SPECIFIC  ROLE  OF  AMINO  ACIDS  IN 
NUTRITION  AND  GROWTH 

That  the  chemical  differences  in  proteins  as  deter- 
mined by  their  amino  acid  content  must  be  of  con- 
siderable significance  in  metabolic  processes  has  been 
understood  in  a  vague  way  for  a  long  time.  As  soon 
as  recognition  was  gained  for  the  view  that  the  prob- 
lems of  nutrition  are  concerned  with  other  factors 

'than  a  mere  sufficiency  of  nitrogen  or  an  adequate 
intake  of  potential  energy  the  problems  of  interme- 
diary metabolism  forced  themselves  upon  the  atten- 
tion of  physiologists  and  led  to  a  thorough  apprecia- 
tion of  the  value  in  nutrition  of  factors  previously 
entirely  overlooked  or  considered  of  little  or  no 
moment. 

Reference  to  the  table  on  p.  22  will  bring  out  clearly 
the  differences  that  exist  between  a  few  of  the  typical 
proteins.  The  most  striking  variations  in  amino  acids 
from    a    quantitative   viewpoint   are    evident.      Such 

■  differences  are  undoubtedly  of  importance  from  a 
nutritional  standpoint,  but  of  much  greater  signifi- 
cance are  the  qualitative  variations.  To  point  out 
briefly  the  most  evident  of  these  it  may  be  seen  that 


NUTRITION  AND  GROWTH  137 

albumin  and  casein  are  glycocoU-free.  Gliadin  from 
wheat  contains  no  glycocoll  and  only  a  trace  of  lysine. 
Zein  from  maize  yields  no  lysine  nor  tryptophane  and 
gelatine  contains  no  cystine,  tyrosine  nor  tryptophane. 
The  first  appreciation  that  qualitative  differences 
in  protein  composition  may  be  of  importance  in  nutri- 
tion was  furnished  by  the  classic  experiments  of  Voit 
and  Munk,  who  showed  that  gelatin  could  not  support 
nitrogen  equilibrium.  The  demonstration  by  Escher 
that  the  addition  of  tyrosine  improved  the  powers  of 
gelatin  in  establishing  nitrogenous  equilibrium  gave 
rise  to  a  series  of  investigations,  the  results  of  which 
have  led  to  a  much  more  complete  understanding  of 
the  problems  intimately  connected  with  metabolism. 
Only  a  few  of  these,  however,  need  be  reviewed  here, 
Kaufman  was  able  to  show  that  when  gelatin  is  fed 
to  man  and  dogs  with  the  addition  of  the  missing 
amino  acids,  tyrosine  and  tryptophane,  nitrogen  equi- 
librium could  be  maintained  for  short  periods  at  least. 
The  work  of  Willcock  and  Hopkins  with  zein,  which 
it  will  be  remembered  is  deficient  in  lysine  and  tryp^ 
tophane,  is  of  great  interest  in  the  present  discussion 
as  it  attacked  the  problem  from  new  viewpoints,  in 
entire  accord  with  the  conceptions  of  the  present.  In 
their  introduction  these  authors  point  out  that:  "We 
are  no  longer  bound  to  Liebig's  view,  or  to  any  modi- 
fication of  it  which  implies  that  the  whole  of  the 
protein  consumed  is  utilized  as  intact  protein :  nor  are 
we  even  compelled  to  assume  that  the  whole  of  what 
is  broken  down  in  the  gut  is  resynthesized  before 


138  THE  AMINO  ACIDS 

utilization.  Protein  products  may  function  in  other 
ways  than  in  the  repair  of  tissues  or  in  supplying 
energy.  It  is  highly  probable  that  the  organism  uses 
them,  in  part,  for  more  specific  and  more  immediate 
needs.  The  discovery  of  substances  absolutely  essen- 
tial to  life,  highly  specific,  and  elaborated  in  special 
organs,  suggests  that  some  part,  at  least,  of  the  pro- 
tein products  set  free  in  the  gut  may  be  used  directly 
by  these  organs  as  precursors  of  such  specific  sub- 
stances. In  adrenaline,  for  instance,  we  have  an  aro- 
matic substance  absolutely  essential  for  the  mainten- 
ance of  life,  and  it  is  probable  that  the  suprarenal 
gland  requires  a  constant  supply  of  some  one  of  the 
aromatic  groups  of  the  protein  molecule  to  serve  as 
an  indispensable  basis  for  the  elaboration  of  adrena- 
line. If  this  be  so,  it  is  certain  that  the  gland  itself 
could  not,  in  starving  animals,  supply  sufficient  of  such 
a  precursor  to  outlast  the  observed  survival  periods. 
Since  adrenaline  must  be  produced  at  all  costs,  the 
required  precursor  must,  in  starvation,  be  obtained  by 
tissue  breakdown  outside  the  gland.  We  may  be  sure, 
however,  that  adrenaline  is  far  from  being  the  only 
substance  elaborated  to  which  such  considerations 
apply.  Similarly,  in  an  animal  upon  a  diet  sufficient 
to  supply  energy,  but  lacking  in  some  essential  group, 
the  minimal  waste  in  the  general  tissues  of  the  body 
will  be  determined  by  the  special  need  of  the  organs  for 
the  missing  group.  On  this  basis  we  have  a  hypoth- 
esis to  account  for  the  special  protein-sparing  prop- 
erties   of    gelatin.      It   shares    with   protein    certain 


NUTRITION  AND  GROWTH  139 

molecular  groupings  needed  to  satisfy  specific  needs, 
and  is  thus  superior  to  fats  and  carbohydrates  as  a 
protein-sparer:  it  lacks,  on  the  other  hand,  certain 
necessary  groupings,  fails  therefore  to  supply  all  such 
needs,  and  thus  cannot  replace  protein." 

These  considerations  served  as  the  basis  for  the* 
experiments  described  by  Willcock  and  Hopkins.  Mice 
kept  under  exactly  similar  conditions  were  fed  with  a 
diet  having  zein  as  its  source  of  nitrogen.  In  certain 
instances  small  quantities  of  tyrosine  or  tryptophane 
were  added  to  the  dietary.  The  results  of  the  influ- 
ence of  such  diets  were  measured  by  the  "survival 
period" — that  is,  th«  period  necessary  to  cause  the 
death  of  the  animal.  In  Fig.  1  is  reproduced  a  dia- 
gram illustrating  very  clearly  the  influence  of  tryp- 
tophane upon  the  survival  period.  With  zein  as  the 
only  nitrogenous  component  of  the  diet  young  miQe 
were  shown  to  be  unable  to  maintain  growth.  Tryp- 
tophane addition  does  not  make  zein  capable  of  main- 
taining growth,  but  does  greatly  prolong  the  survival 
period.  In  Fig.  1  the  survival  periods  of  mice  fed 
upon  zein  alone  are  not  given,  for  they  were  identical 
with  those  obtained  with  mice  fed  zein  plus  tyrosine. 
Although  added  tyrosine  exerted  no  influence  upon 
the  survival  period,  it  must  not  be  inferred  that  this 
amino  acid  is  without  specific  effect  on  metabolism: 
it  Evidently  played  little  role  here  because  zein  fed 
supplied  sufficient  tyrosine,  hence  an  excess  was  with- 
out special  influence.  In  reality  tyrosine  was  added  as 
a  control  to  tryptophane  addition,  in  order  to  determine 


A-       8       12       16      20     24      28     32     36    ^10     ^44     ^8    da 

Figure  1.  The  thick  lines  show  the  survival  periods  (in  days)  of  twenty-o 
individual  mice  upon  the  zein  diet  with  tyrosine  added.  The  thin  lines  show  t 
same  for  nineteen  mice  upon  the  zein  diet  with  tryptophane  added.  [From 
Journal  of  Physiology^  volume  35.] 


NUTRITION  AND  GROWTH  141 

whether  addition  of  any  amino  acid  would  produce 
an  effect,  and  hence,  therefore,  to  find  out  directly  the 
specific  action  of  tryptophane. 

A  prominent  feature  in  connection  with  the  mice 
given  zein  alone  was  a  condition  of  torpor;  the  mice 
were  very  inactive  and  made  no  movement  when 
handled  or  touched,  the  ears,  feet,  and  tail  were  cold, 
the  coat  was  glairy  and  the  eyes  were  half-closed. 
Those  fed  tryptophane  With  zein  showed  a  strikingly 
different  behavior,  being  active  and  apparently  healthy 
even  up  to  the  end  of  life.  In  both  instances  death 
was  not  caused  by  a  lack  of  food  intake,  as  all  animals 
gave  evidence  of  appetite.  Quantitatively,  sufficient 
food  was  received  but  qualitatively  something  essen- 
tial to  life  was  lacking.  It  is  possible  that  had  lysine, 
the  other  amino  acid  lacking  in  zein,  been  fed  also, 
even  better  results  would  have  been  obtained.  Tryp- 
tophane  undoubtedly  is  essential  for  the  maintenance 
of  life,  although  the  specific  role  it  plays  has  not  yet 
been  determined.  As  the  authors  mentioned  above 
point  out,  "If  it  [tryptophane]  serves  as  a  basis  for 
the  elaboration  of  a  substance  absolutely  necessary 
for  life — something,  for  instance,  of  an  importance 
equal  to  that  of  adrenaline — then,  in  starvation,  or 
when  it  is  absent  from  the  diet,  a  supply  is  likely  to 
be  maintained  from  the  tissue-proteins,  the  demand 
for  it  would  become  one  of  the  factors  determining 
tissue  breakdown.  In  the  case  of  young  animals  which 
directly  benefit  from  the  addition  of  a  protein  con- 
stituent otherwise  absent  from  their  diet,  to  the  extent 


142  THE  AMINO  ACIDS 

of  a  well-nigh  doubled  life,  and  lose,  instead  of  gaining, 
weight,  the  utilization  of  the  constituent  would  seem  to 
be  of  some  direct  and  specific  nature."  These  words 
give  the  first  definite  suggestion  that  individual  amino 
acids  may  play  a  specific  role  in  the  maintenance  of 
nutritional  rhythm. 

The  failure  of  zein  as  a  suitable  source  for  the 
essential  nitrogen  requirement  leads  to  the  query 
whether  any  single  protein  will  suffice  in  this  respect. 
Attempts  to  answer  this  question  have  been  many  and 
it  is  only  recently  that  a  satisfactory  positive  reply  has 
been  given.  In  many  of  the  older  experiments  lack  of 
success  has  been  attributed  to  various  factors  other 
than  the  character  of  the  protein,  and  where  appar- 
ently successful  results  have  been  obtained  criticism 
has  been  pertinent  in  that,  in  most  instances,  the  protein 
or  proteins  employed  have  not  been  free  from  impuri- 
ties. The  general  impression  gained  from  this  type 
of  investigation  has  been  that  sooner  or  later  animals 
die  when  kept  for  a  prolonged  period  upon  a  con- 
stant diet  even  though  an  abundance  of  energy 
producing  material  may  be  present.  A  so-called 
"pure"  diet  has  been  deemed  impracticable.  Lunin,  one 
of  the  early  investigators  of  the  problem,  fed  mice 
with  mixtures  of  casein,  fat,  cane  sugar,  and  milk 
ash.  On  this  artificial  diet  death  occurred  in  from 
twenty  to  thirty  days,  a  survival  period  greater  than 
when  the  ash  of  milk  was  omitted.  Mice  fed  dried 
milk  were  alive  at  the  end  of  two  months.  Hall  with 
mice   and    Steinitz   with   dogs   obtained   comparable 


NUTRITION  AND  GROWTH  143 

results  when  a  similar  form  of  dietary  was  used.  By 
considerable  variation  in  the  non-nitrogenous  portion 
of  the  food  Rohmann  showed  that  mice  will  thrive 
for  weeks.  A  criticism  of  these  experiments  is  that 
the  range  of  variation  in  the  make-up  of  the  dietary 
resulted  really  in  furnishing  the  animals  an  ordinary 
mixed  diet.  The  experiments  of  Jacob  with  pigeons, 
of  Falta  and  Noeggerath,  and  of  Knapp  with  rats 
demonstrated  that  variety  in  the  dietary  undoubtedly 
tends  toward  prolongation  of  life  but  that  death 
eventually  ensues. 

After  experiencing  many  failures,  Osborne  and 
Mendel  have  succeeded  in  maintaining  white  rats  for 
long  periods  of  time  upon  single,  pure,  isolated  pro- 
teins, growth  also  being  at  a  normal  rate.  They 
attributed  their  success  to  the  addition  to  the  dietary 
of  what  they  term  "protein-free  milk."  This  is  pre- 
pared by  removing  the  protein  and  fat  from  milk, 
leaving  the  milk  sugar,  inorganic  salts  and  the  un- 
known components.  "Protein-free  milk"  always  con- 
tains very  small  quantities  of  protein  but  not  enough 
to  support  life.  They  have  also  demonstrated  that 
by  artificially  imitating  the  composition  of  "protein- 
free  milk"  by  union  of  the  various  ions  fairly  success- 
ful results  have  been  obtained.  It  is  therefore  pos- 
sible to  construct  a  dietary  in  such  a  manner  from 
purified  isolated  foodstuffs  and  artificial  salt  mixtures 
that  animals  may  not  only  be  maintained  but  normal 
growth  may  also  be  induced. 

In  their  work,  Osborne  and  Mendel  differentiate 


144  THE  AMINO  ACIDS 

sharply  between  a  maintenance  diet  and  one  capable 
of  promoting  growth.  They  have  shown,  for  example, 
that  a  young  animal  may  be  maintained  on  a  certain 
diet  indefinitely  without  manifesting  any  tendency  to 
grow.  From  the  work  of  Donaldson  it  has  been  dem- 
onstrated that  the  life  span  of  the  white  rat  is  about 
three  years.  Sexual  maturity  is  reached  in  sixty 
days.  The  first  year  of  life  for  the  rat  corresponds 
to  the  first  thirty  years  of  human  life,  and  the  curve 
of  growth  for  this  period  is  reproduced  below.  Fig.  2. 
As  an  illustration  of  the  influence  of  an  isolated 
protein,  casein  ( fed  with  .  starch,  sugar,  agar,  lard, 
and  a  salt  mixture),  the  chart.  Fig.  3,  is  shown.  It 
is  evident  that  casein  as  the  sole  source  of  nitrogen 
was  apparently  incapable  of  allowing  normal  growth 
in  a  young  rat  during  a  period  of  forty-six  days.  In 
other  words,  stunting  occurred.  In  period  2,  casein 
and  sugar  were  replaced  by  milk.  Growth  was 
resumed.  The  influence  of  changing  the  salt  mixture 
content  of  the  food  intake  is  quite  evident  in  periods 

3,  4,  and  5.  The  ability  of  milk  to  furnish  the  necessary 
nitrogen  requirement  is  well  shown  in  the  chart,  Fig. 

4,  the  curve  obtained  being  to  all  intents  and  purposes 
identical  with  the  normal  growth  curve. 

If  to  the  casein  diet  "protein-free  milk"  is  added, 
instead  of  whole  milk  replacing  casein,  normal  condi- 
tions obtain  as  is  well  illustrated  in  the  chart.  Fig.  5. 

Casein  alone  was  found  to  be  unable  to  support 
growth.  In  Fig.  6  is  shown  a  curve  in  which,  during 
period  2,  casein  was  the  only  source  of  protein  and 


300 


280 


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Figure  2  shows  average  normal  rates  of  growth  of  male 

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NUTRITION  AND  GROWTH 


145 


as  a  result  a  decline  set  in,  which  could  not  be  checked 
by  doubling  the  percentage  of  casein  in  the  diet.  That 
lack  of  protein  can  not  account  for  the  decline  is  well 
shown  in  period  4,  during  which  the  original  amount 


t«rm»nal6d 


Figure  4.    Growth  Curve  with 
Milk  Diet. 


of  casein  was  replaced  and  "protein-free  milk"  was 
also  added.  An  immediate  response  in  appetite  was 
evidenced  and  speedy  recuperation  and  growth  were  in 
order.  This  experiment  demonstrates  that  a  rat  unable 
to  maintain  itself  on  an  isolated  protein  may  be  caused 


146 


THE  AMINO  ACIDS 


to  speedily  resume  a  normal  condition  by  the  addition 
to  the  diet  of  "protein-free  milk." 

From  these  and  many  similar  results  it  is  apparent 
that  if  suitable  non-protein  constituents  of  the  dietary 


20 


40 


60  80  too         120 

Days 


Figure  5.    Maintenance  on  Casein  and  Growth 
AFTER  Addition  of  Protein-free  Milk. 


are  supplied,  such  as  are  furnished  by  "protein-free 
milk"  maintenance  and  growth  in  white  rats  may  be 
normal.  Emphasis  should  therefore  be  laid  upon  the 
importance  of  the  role  played  by  the  accessory  food- 
stuffs, as  contained  in  "protein-free  milk"  the  nature 


SUJ9J0 


148  THE  AMINO  ACIDS 

of  which  remains  obscure.  It  is  also  evident  that  the 
establishment  of  a  satisfactory  non-protein  dietary 
aifords  an  opportunity  for  the  study  of  any  specific 
influence  which  a  peculiar  type  of  protein,  or  one  with 
an  unusual  type  of  internal  structure,  may  exert  in 
nutrition. 

In  addition  to  casein  Osborne  and  Mendel  have 
demonstrated  that  perfectly  satisfactory  results  may 
be  yielded  when  other  types  of  pure  proteins  are 
employed,  a  single  one  sufficing  for  all  the  nitrogen 
requirements  of  white  rats.  Thus,  adequate  growth 
has  been  secured  with  lactalbumin  from  cow's  milk, 
ovalbumin  from  hen's  egg,  ovovitellin  from  hen's  egg, 
edestin  from  hemp  seed,  cannabin  from  hemp  seed,  glu- 
tenin  from  wheat,  glycinin  from  the  soy  bean,  globulin 
from  squash  seed,  globulin  from  cotton  seed,  excelsin 
from  Brazil  nut,  and  glutelin  from  maize. 

Taking  advantage  of  the  opportunity  afforded  them, 
the  above  mentioned  authors  have  studied  the  influ- 
ence which  a  peculiar  protein,  for  example,  one  lack- 
ing one  or  more  important  amino  acid,  may  exert  in 
nutritional  processes.  It  soon  became  evident  that  all 
proteins  do  not  promote  growth  under  otherwise 
favorable  conditions.  Gliadins  .  of  rye  and  wheat, 
which  are  deficient  in  glycocoll  and  lysine  and  on  the 
other  hand  are  very  rich  in  glutamic  acid,  and  hordein 
of  barley,  which  closely  resembles  gliadin  in  chemical 
constitution,  are  capable  of  giving  maintenance,  but 
fail  to  induce  growth.  A  condition  of  stunting  is 
brought  about,  old  animals  retaining  the  characteris- 


NUTRITION  AND  GROWTH 


149 


tics  of  well-nourished  young  rats.  In  Fig.  7  are  re- 
produced curves  which  show  the  failure  of  a  rat  to 
present  normal  growth  on  a  diet  containing  protein- 
free    milk    and   gliadin   as    the   only    protein.      The 


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Figure  7.    Failure  of  Growth  on 
Gliadin  plus  Protein-free  Milk. 

frontispiece  shows  the  photograph  of  this  rat  (B) 
and  as  a  contrast  that  of  a  rat  (A)  of  the  same 
age  presenting  normal  growth,  together  with  a  pho- 
tograph of  a  rat  (C)  of  the  same  weight  as  (B)  but 
much  younger.    This  stimting  is  apparently  a  method 


150  THE  AMINO  ACIDS 

which  may  be  employed  for  the  attainment  of  a  type 
of  animal  infantilism.  In  connection  with  the  sub- 
ject of  stunting  it  became  of  interest  to  determine 
whether  this  condition  would  remain  permanent  under 
all  circumstances  or  whether  a  return  to  a  diet  con- 
taining a  more  typical  protein  than  gliadin  would  also 
cause  a  resumption  of  growth.  Fig.  8  shows  the 
slight  growth  of  a  young  white  rat  during  276  days 
of  gliadin  feeding.  That  the  capacity  to  grow  had 
not  been  lost,  but  was  merely  inhibited,  may  be  seen 
in  the  second  part  of  the  curve  in  which  milk  food 
replaced  the  gliadin.  At  the  beginning  of  the  milk 
food  diet  the  rat  was  314  days  old,  an  age  at  which 
rats  usually  show  very  little  growth.  Fertility  is  not 
impaired  by  the  act  of  stunting,  as  may  be  seen  from 
the  curve  in  Fig.  9,  for  this  rat,  after  a  period  of  154 
days  with  gliadin  as  its  protein  supply,  was  mated 
and  produced  four  young,  which  were  suckled  during 
the  first  month  of  their  existence  by  the  mother  who 
was  still  maintained  upon  a  gliadin  diet.  These  young 
rats  presented  normal  growth  curves  during  this 
period.  When  a  month  old,  three  of  the  young  ani- 
mals were  removed  from  the  mother  and  kept  upon 
diets  of  casein,  edestin,  and  milk  food.  All  showed 
normal  curves  of  growth.  The  fourth  young  rat, 
kept  with  the  mother  began  to  exhibit  a  failure  to 
grow  as  soon  as  forced  to  depend  upon  the  gliadin 
food  mixture.  Inasmuch  as  casein,  which  has  been 
proved  to  be  efficient  as  a  source  of  nitrogen  for  both 
maintenance    and    growth,    is    lacking    in    glycocoll. 


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Figure  8.  This  figure  shows  failure  of  rat  to  make  more  than  slight  gn 
at  a  normal  rate  after  276  days  of  stunting.  At  this  time  the  rat  was  314  day 
Biological  Che^nistry ^  volume  12.] 


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)n  a  diet  containing  gliadin  as  the  sole  protein,  and  capacity  to  resume  growth 
'^■"  age  at  which  rats  normally  grow  very  little  more.     [From  the y6>«r«a/ ^ 


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Figure  9  shows  maintenance  and  fertility  on  a  diet  containing  gliadin  as  its 
sole  protein.  After  154  days  this  rat  was  paired,  four  young  being  the  result  of 
the  mating.     [From  the  Journal  of  Biological  Chemistry^  volume  12.] 


152  THE  AMINO  ACIDS 

whereas  gliadin  is  deficient  in  glycocoU  and  lysine  and 
fails  to  promote  growth,  it  is  reaso^abfe  to  assume 
that  the  low  content  of  lysine  in  gliadm  is  responsible 
for  the  failure  of  white  rats  to  grow.  On  the  other 
hand,  lysine  is  apparently  not  essential  for  mere  main- 
tenance. Another  conclusion  which  may  be  drawn 
from  these  experiments  is  that  the  organism  is  unable 
to  synthesize  lysine,  although  glycocoll  may  be  syn- 
thesized with  apparent  ease,  as  has  been  shown  in 
previous  pages  of  this  book.  Growth  means  the  for- 
mation of  new  tissues  and  in  the  absence  of  sufficient 
lysine  the  construction  of  new  tissue  does  not  occur 
readily,  or  at  least  proceed  at  the  normal  rate.  The 
inference  that  lysine  is  concerned  with  the  function 
of  growth  may  be  tested  from  another  viewpoint.  If 
the  animals  fed  with  gliadin,  lacking  in  lysine,  show 
a  failure  to  grow  the  addition  of  lysine  to  gliadin 
should  be  followed  by  a  resumption  of  normal  growth. 
Such  trials  have  been  made  by  Osborne  and  Mendel 
and  the  results  obtained  are  most  strikingly  seen  in 
the  following  curves.  [See  Fig.  10.]  Failure  to 
grow  on  gliadin  as  the  sole  protein  is  first  shown  in 
the  curves  followed  by  a  period  of  growth  when 
lysine  was  added  to  the  diet.  The  subsequent  with- 
drawal of  the  lysine  is  followed  in  each  instance  by 
a  cessation  of  growth.  If  lysine  is  added  again  growth 
is  again  resumed  at  a  normal,  to  cease  again  when 
lysine  is  taken  away.  These  results  lead  to  the  con- 
clusion that  lysine  is  indispensable  for  the  functions 
of  growth.     Data  collected  by  Osborne  and  Mendel 


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Figure  11.  Experiments  with  Zein.  Neither  growth  normain- 
tenance  can  be  secured  when  zein  is  the  sole  protein  in  the  dietary.  [From 
ih&  Journal  of  Biological  Chemistry^  volume  17.] 


NUTRITION  AND  GROWTH  155 

reveal  the  "teleologically  interesting  fact  .  .  .  that 
those  proteins,  Hke  casein,  lactalbumin,  and  egg  viteUin, 
which  are  in  nature  concerned  with  the  growth  of 
animals,  all  show  a  relatively  high  content  of  lysine." 
The  experience  of  these  investigators  with  zein, 
which  lacks  glycocoU,  tryptophane  and  lysine,  has 
brought  to  light  the  fact  that  tryptophane  is  undoubt- 
edly essential  for  maintenance  and  emphasizes  anew 
the  significance  of  lysine  as  a  growth  promoting  sub- 
stance. One  may  also  assume  that  a  little  lysine  is 
necessary  for  maintenance  and  this  is  ordinarily  sup- 
plied in  sufficient  amount  by  the  traces  in  gliadin  or 
(in  the  zein  and  tryptophane  experiments)  by  traces 
in  protein-free  milk  protein  or  from  the  tissues  them- 
selves. In  an  earlier  portion  of  this  chapter  were 
pointed  out  in  some  detail  the  experiments  of  Will- 
cock  and  Hopkins  with  zein,  with  and  without  addi- 
tion of  tryptophane.  They  found  that  zein  as  the 
only  protein  in  the  dietary  cannot  maintain  growth  in 
the  young  animal  nor  even  support  life.  The  addi- 
tion of  tryptophane  resulted  in  prolonging  life  with- 
out causing  a  resumption  of  the  growth  impulse. 
The  outcome  of  the  work  of  Osborne  and  Mendel 
with  zein  alone  is  best  shown  in  the  chart,  Fig.  11. 
The  large  number  of  experiments  shown  here  yielded 
concordant  results  and  show  that  neither  maintenance 
nor  growth  can  be  secured  when  zein  is  the  only  pro- 
tein ingested.  When  tryptophane  is  added  to  the  zein 
food  mixture,  maintenance  of  body  weight  follows,  as 
may  be  seen  from  Fig.  12.    Addition  of  both  trypto- 


156 


THE  AMINO  ACIDS 


Days 


Figure  12.  Indispensability  of  Tryptophane  for  Main- 
tenance IN  Nutrition.  These  experiments  should  be  contrasted 
with  the  failure  of  maintenance  on  zein-food  alone,  shown  in  figure 
11.     [From  the  Journal  of  Biological  Chemistry,  volume  17.] 

pliane  and  lysine  results  in  the  establishment  of  perfect 
maintenance  and  growth.  [See  Fig.  13.]  It  may  be 
inferred  from  these  experiments  then  that  tryptophane 
is  indispensable  for  maintenance  in  nutrition  and  that 
the  animal  organism  does  not  possess  the  ability  to 
synthesize  this  amino  acid.  That  lysine  cannot  replace 
tryptophane  in  the  establishment  of  the  condition  of 


Days 


Figure  13.  Growth  on  Foods  Containing  Zein+Tryptophanb 
+LYSINE.  The  growth  obtained  on  this  diet  may  be  contrasted  with 
maintenance  without  growth  in  the  absence  of  the  lysine  (see  Figure  12) 
2.ndi  failure  to  be  maintained  in  the  absence  of  both  lysine  and  tryptophans 
(Figure  11),  thus  demonstrating  the  role  of  these  amino  acids  in  growth 
and  maintenance  respectively.  That  lysine  cannot  replace  tryptophane  in 
maintenance  is  shown  by  Rat  1900.  [From  the  Journal  of  Biological 
Chemistry^  volume  17.] 


158  THE  AMINO  ACIDS 

maintenance,  may  be  seen  from  the  chart,  Fig.  13. 
Rat  1900. 

Investigation  of  this  type  into  the  biochemical  de- 
portment of  the  protein  cleavage  products  will  un- 
doubtedly lead  ultimately  to  the  assignment  of  more 
or  less  specific  functions  to  the  various  amino  acids, 
and  hence  will  indirectly  indicate  the  relative  efficiency 
of  this  or  that  protein  in  bringing  about  a  desired 
result  in  nutrition. 


References  to  Literature 

Mendel:   Nutrition   and   Growth :   Journal  of  the  American 
Medical  Association.    1915,  64,  p.  1539. 

Osborne  and  Mendel:   Journal   Biological   Chemistry.     1914, 
ir,  p.  325. 

Willcock  and  Hopkins:  Journal  of  Physiology.    1906,  35,  p.  88. 


INDEX 


INDEX 


Absorption, 
from  intestine,  48. 
from  large  intestine,  56. 
from  stomach,  46,  47. 
of  amino  acids,  47,  54,  56. 
of  amino  acids  by  rectum, 

131. 
of  amino  acids  by  tissues, 

of  fat,  53. 

of    proteoses    and    pep- 
tones, 52,  56. 

of  putrefactive  products, 
56. 

of  undigested  protein,  48. 
Accessory  foodstuffs,  146. 
Acetic  acid,  101. 
Acetone,  101. 
Acid,  acetic,  101. 

aspartic,  16. 

caseinic,  18. 

glutamic,  16. 

hippuric,  108. 

homogentisic,  106,  115. 

isovaleric,  100. 

lactic.  111,  112,  113. 

nucleic,  7. 

uric,  93. 
Acids,  amino,  12. 

diamine,  21. 

hydroxy,  38. 


Acids,  ketone,  100. 
monoamine,  21. 
Adrenaline,  see  also  Epine- 
phrine, 138,  141. 
Alanine,  13. 
amounts  of,   in  proteins, 

22,  23. 
dextrose  formation  from, 

111. 
formation  from  glycogen, 

113. 
in  blood,  55. 
Albuminates,  9. 
Albuminoids,  6. 
Albumins,  4,  5. 
Alcohol-soluble  proteins,  4, 

5. 
Alkaptonuria,  114. 
Amides,  as  foodstuffs,  131. 
Amines,  40. 

fate  of,  57. 
Amino   acid,   definition   of, 
12. 
metabolism,  anomalies  of, 
114. 
Amino  acids,  12. 
absorption  of,  54,  56,  Id. 
absorption  of,  by  rectum, 

131. 
action  of  intestinal  bac- 
teria upon,  38. 


162 


INDEX 


Amino  acids,  as  foodstuffs, 
126,  131. 

as  functional  test  of  liver, 
71. 

as  protein  sparers,  127. 

content  of,  in  tissues,  11, 
78,  79. 

deficiencies  of,  in  gliadin, 
148. 

deficiencies  of,  in  zein, 
155. 

description  of,  12. 

fate  of,  in  tissues,  76. 

formation  of  carbohy- 
drates from,  109. 

formation   of,   from   car- 

.  bohydrates,  113. 

formation  of,  in  gastric 
digestion,  31. 

formation  of,  in  intestinal 
digestion,  35. 

formulas  of,  12. 

further  fate  of,  99. 

in  blood,  55,  IZ,  75,  78. 

in  digestion,  28,  29. 

in  duodenal  contents,  35. 

in  intermediary  metabo- 
lism, 78. 

in  maintenance  and 
growth,  148. 

mono,  21. 

quantitative  yields  from 
proteins,  22. 

relationship  of  different, 
19. 


Amino  acids,  relation  of,  to 
specific  dynamic  action, 
123. 

specific  role  of,  in  nutri- 
tion and  growth,  136. 

synthesis  of,  107,  132,  134. 

synthesis    of,    to   protein, 

n. 

Ammonia,  93. 

amounts   of,   in   proteins, 

22,  23. 
in  intestinal  putrefaction, 
39. 
Ammonium  salts,  as   food- 
stuffs, 131. 
Amounts    of    amino    acids 
yielded  by  protein^,  22. 
Anabolism,  81. 
Animal  infantilism,  150. 
Anomalies    of    amino    acid 

metabolism,   114. 
Arginase,  71. 
Arginine,    17. 
amounts   of,   in   proteins, 

22,  23. 
catabolism  of,  106. 
dextrose  formation  from, 

112. 
fate   of,    in   putrefaction, 

44. 
in  blood,  55. 
urea  from,  71. 
Arbacin,  6. 

Artificial  foodstuffs,  value 
of,  in  nutrition,  143. 


INDEX 


163 


Artificial      production      of 

foodstuffs,  130. 
Asparagine,  food  value  of, 

132. 
Aspartic  acid,  16. 
amounts   of,   in   proteins, 

22,  23. 
dextrose  formation  from, 

112. 
in  blood,  55. 
Bacterial      digestion      and 

amino  acids,  36. 
j3-iminazolylethylamine,   43. 
jS-oxybutyric  acid,  102. 
Blood,   amino   acids  in,   55, 
73,  75. 
fate  of  amino  acids  in,  76. 
non-coagulable       protein, 

60. 
non-protein    nitrogen    of, 

55. 
proteose  of,  50. 
proteoses  and  peptones  in, 

59. 
seromucoid  in,  61. 

Cadaverine,  43,  147. 
Cannabin,  growth  with,  148. 
Carbohydrate,  formation  of 

amino  acids  from,  113. 
formation  of,  from  amino 

acids,  109. 
"Carbon  moiety"  of  protein, 

110. 
Casein,   as   sole  protein   of 

diet,  144. 


Caseinic  acid,  18. 

amount  in  casein,  22. 
Catabolism,  81. 

of  amino  acids,  99. 
Circulating  protein,  84. 
Classification  of  proteins,  3. 
Clupein,  6. 

Coagulated  proteins,  9. 
Coagulation  of  protein,  2. 
Colloids,  2. 

Conjugated  proteins,  4,  7. 
Creatinine,  92. 
Cresol,  38. 
Cystine,  15. 

absence  of,  in  gelatin,  137. 

amounts   of,   in   proteins, 
22,  23. 

excretion  of,   in   cystinu- 
ria,  117. 
Cystinuria,  116. 

Deamination,  70,  71,  72,  99, 
100,  101. 

Derived  proteins,  4,  8. 

Dextrose,  formation  of, 
from  amino  acids.  111, 
112. 

Diacetic  acid,  104. 

Diamines,  43,  117. 

Diamino  acids,  21. 

Diaminuria,  116. 

Diet,  variety  in,  143. 

Digestion,  a  hydrolytic  pro- 
cess, 29. 
and  amino  acids,  28. 


164 


INDEX 


Edestin,  growth  with,  148. 
Endogenous  metabolism,  95. 
Enzymes,     in    blood    after 

protein    injections,    51, 

61. 
in    protein    synthesis,    67, 

69. 
Epinephrine,        see        also 

Adrenaline,  41. 
Erepsin,  34. 
Ethereal  sulphates,  94. 
Excelsin,  growth  with,  148. 
Excretion    of    putrefactive 

products,  56. 
Exogenous  metabolism,  95. 

Fertility,  and  stunting,  150. 
influence    of    gliadin    on, 
150. 
Fibrin,  9. 
Fibrinogen,  9. 
Foodstuffs,  amides  as,  131. 
amino  acids  as,  126,  131. 
ammonium  salts  as,  131. 
artificial    production     of, 

130. 
value  of  artificial,  in  nu- 
trition, 143. 

Gaduhiston,  6. 

Gastric     digestion,     impor- 
tance of,  32. 
products  of,  30,  31,  32. 
relation  of,  to  amino  acid 
formation,  32, 
Gelatin,  absence  of  certain 
amino  acids  in,  137. 


Gelatin,  as  a  protein  sparer, 
138. 
nutritive  value  of,  137. 
Gliadin,  5. 
deficiency  of  amino  acids 

in,  148. 
influence  of,  on  fertility, 

150. 
influence  of,   on  growth, 

150. 
yield  of  lysine  and  glyco- 
coll,  137. 
Globin,  6. 
Globulins,  4,  5. 

growth  with,  148. 
Glucosamine,  7. 
Glucoproteins,  7. 
Glutamic  acid,  16. 
amounts   of,   in  proteins, 

22,  23. 
dextrose  formation  from, 

112. 
in  blood,  55. 
in  gliadin,  148. 
Glutelin,  growth  with,  148. 
Glutelins,  4,  5. 
Glutenin,  5. 

growth  with,  148. 
Glycinin,  growth  with,  148. 
Glycocoll,  12. 
absence    of,    in    gliadins, 

137,  148. 
amounts   of,  in  proteins, 

22,  23. 
dextrose  formation  from, 
111. 


INDEX 


165 


Glycocoll,  in  blood,  55. 

synthesis  of,  108. 
Glycogen,  109. 

formation      of       alanine 
from,  113. 
Growth,    and    maintenance, 
143. 
influence  of  lysine  upon, 

152,  155,  156. 
influence    of    milk    food 

upon,  144. 
influence   of    tryptophane 

upon,  155,  156. 
influence    of    zein    upon, 

139,  155. 
specific     role     of     amino 
acids   in  nutrition  and, 
136. 
with  various  proteins,  148. 
Heat  production,   and   me- 
tabolism, 89. 
Hemocyanin,  8. 
Hemoglobins,  8. 
Heredity,    in    alkaptonuria, 
115. 
in  cystinuria,  117. 
Heterocyclic  compounds,  21. 
Hippuric  acid,  108. 
Histamine,  43. 
Histidine,  17. 
amounts   of,   in   proteins, 

22,  23. 
fate   of,   in   putrefaction, 

42,  43. 
in  blood,  55. 
Histones,  6. 


Homogentisic  acid,  106. 
relation    of,    to    tyrosine 
and  phenylalanine,   115. 
Hydrolysis,  of  protein,  11. 
Hydroxy  acids,  38,  39. 

Indole,  39. 

ethylamine,  42. 
Inorganic  sulphates,  93. 
Intestinal   digestion,   33. 
relation  of,  to  amino  acid 
formation,  34. 
Intestinal    work,     influence 
of,  in  specific  dynamic 
action,  112. 
Isoamylamine,  42. 
Isoleucine,  14. 
amounts   of,   in   proteins, 
22,  23. 
Isovaleric  acid,  100. 

Ketone  acids,  100. 

Lactalbumin,    growth   with, 

148. 
Lactic  acid.  111,  112,  113. 
Lecithins,  8. 
Lecithoproteins,  8, 
Leucine,  13. 
amounts   of,   in   proteins, 

22,  23. 
catabolism  of,   100. 
fate   of,    in   putrefaction, 

42. 
in  blood,  55. 
Leucocytes,  role  of,  in  pro- 
tein synthesis,  66. 


166 


INDEX 


Liver,  in  amino  acid  metab- 
olism, 71. 

role  of,  in  protein  synthe- 
sis, 129. 
Lysine,  16. 

absence  of,  in  zein,  137. 

amounts   of,   in   proteins, 
22,  23. 

inability  of  body  to  syn- 
thesize, 152. 

in  blood,  55. 

influence   of,   on  growth, 
152,  155,  156. 

in  gliadin,  137,  148. 

in  maintenance,  156. 

Maintenance,    and    growth, 
143. 
influence  of  lysine  upon, 

156. 
influence    of    tryptophane 

upon,  155,  156. 
influence    of    zein    upon, 
155. 
Metabolism,  81. 
and  heat  production,  89. 
of  amino  acids,  99. 
of  plethora,  124. 
Metaproteins,  9. 
Milk,  and  growth,  144,  145. 
food,     influence     of,     on 

stunting,   150. 
protein-free,  143,  144,  145, 
149,  155. 
Monoamino  acids,  21. 
Mucoids,  in  blood,  60,  61. 


Neutral  sulphur,  93. 
Nitrogen,  equilibrium,  97. 

form  needed  by  body,  1. 

in  protein,  1,  2. 

in  tissue  formation,  96. 
Norleucine,  14. 
Nucleic  acid,  7. 
Nucleoproteins,  7. 
Nutrition,    specific    role   of 
amino  acids  in,  136. 

Occurrence,  and  character- 
istics of  proteins,  5. 

Organized  protein,  84. 

Ornithine,  44. 
dextrose  formation  from, 
112. 

Ovalbumin,     growth     with, 
148. 

Ovovitellin,     growth     with, 
148. 

Oxidative   deamination,   99. 

Oxyproline,  18. 
amounts  of,   in  proteins, 
22,  23. 

p.oxyphenylethylamine,  40. 
Parenteral   introduction   of 

protein,  fate  of,  49. 
Peptides,  10,  26. 
Peptone,   action   of  erepsin 

upon,  34. 
in  gastric  digestion,  31. 
Peptones,  10. 
in  intestinal  putrefaction, 

37. 


INDEX 


167 


Phenol,  38. 
Phenylalanine,  14. 
amounts   of,   in   proteins, 

22,  23. 
catabolism  of,  103. 
relation  of,  to  homogen- 
tisic  acid,  115,  116. 
Phosphoproteins,  7. 
Plastein  formation,  68. 
Plastic  foods,  84. 
Polypeptides,  10,  25. 
action   of   enzymes   upon, 

26. 
value   of,   in   amino   acid 
mixtures,  129. 
Proline,  17. 
amounts   of,   in   proteins, 

22,  23. 
dextrose  formation  from, 

112. 
in  blood,  55. 
Protamines,  6. 
Proteans,  8. 

Protein,  action  of  enzymes 
in  synthesis  of,  67,  69. 
as  a  complex  polypeptide, 

26. 
definition  of,  2. 
fate  of  ingested,  58. 
free  milk,   143,   144,   145, 

146,  149,  155. 
tnolecular  weight  of,  3. 
molecule,  2. 

molecule,  structure  of,  24. 
metabolism,    theories    of, 
81. 


Protein,  regeneration,  place 

of,  62. 
sparers,  127,  132,  133,  138. 
synthesis,     by     intestinal 

bacteria,  133. 
synthesis,  by  intestine,  62. 
synthesis,      from     amino 

acids,  by  tissues,  78. 
synthesis,   role   of   leuco- 
cytes in,  66. 
synthesis,  role  of  liver  in, 

129. 
Proteins,  and  growth,  148. 
as  colloids,  2. 
classification  of,  3. 
characteristics  of,  5. 
composition  of,  2. 
conjugated,  4,  7. 
crystallization  of,  2. 
derived,  4,  8. 
influence  of,  on  plane  of 

polarized  light,  3. 
occurrence  of,  5. 
quantities  of  amino  acids 

yielded  by,  22. 
simple,  4,  5. 
specific     dynamic     action 

of,  120. 
Proteose,  in  blood,  50. 
Proteoses,  9,  10. 
action  of  erepsin  on,  34. 
and  peptones,  in  blood,  59. 
in  gastric  digestion,  31. 
in  intestinal  putrefaction, 

37. 
Ptomaines^  117. 


168 


INDEX 


"Pure"  diets,  142. 
Purine  bases,  7. 
Putrefaction,  fate  of  argin- 
ine  in,  44. 

fate   of   histidine   in,   42, 
43. 

fate  of  leucine  in,  42. 

fate  of  tryptophane  in,  39. 

fate  of  tyrosine  in,  38. 

formation  of  ammonia  in, 
39. 

hydroxy  acids  in,  38. 

nature  of,  36. 

products  of  intestinal,  38. 

proteoses  and  peptones  in, 

Putrescine,  43,  117. 
Pyrimidine  bases,  7. 

Rate  of  blood  flow,  IZ. 
Relationship     of     different 

amino  acids,  19. 
"Residual       nitrogen"       of 

blood,  74. 
Respiratory  foods,  84. 

Salmin,  6. 
Scombin,  6. 
Scombron,  6. 
Serine,  15. 
amounts   of,   in   proteins, 

22,  23. 
dextrose  formation  from, 
112. 
Seromucoid,  61. 


Simple  proteins,  4,  5. 

Skatole,  39. 

Specific  dynamic  action,  90, 

120. 
Starvation,  amino  acids  in 

blood  during,  78. 
Structure  of  protein  mole- 
cule, 24. 
Stunting,  144,  149,  150. 
influence    of    milk    food 

upon,  150. 
Sturin,  6. 
Survival    period,    influence 

of  zein  upon,  139,  140, 

141. 
Synthesis   of   amino   acids, 

107. 

Theories    of    fate    of    in- 
gested protein,  58. 
Theories  of  protein  metab- 
olism, 81. 
Theories  of  protein  regen- 
eration, 62. 
Transformations   of   amino 

acids  in  body,  99. 
Tryptophane,  18. 
absence  of,  in  certain  pro- 
teins, 24,  137. 
amounts   of,   in   proteins, 

22,  23. 
catabolism  of,  106. 
importance   of,    for   life, 

141. 
importance   of,   in   nutri- 
tion, 130. 


INDEX 


169 


Tryptophane,     inability     of 
body  to  synthesize,  156. 

influence  of,  on  mainte- 
nance and  growth,  155, 
156. 

value  of,  in  gelatin  feed- 
ing, 137. 

value  of,  in  zein  feeding, 
139,  140,  141. 
Tyramine,  41. 
Tyrosine,  14. 

absence  of,  in  gelatin,  23, 
137. 

amounts  of,  in  proteins, 
22,  23. 

catabolism  of,  105. 

relation  of,  to  homogen- 
tisic  acid,  115,  116. 

value  of,  in  gelatin  feed- 
ing, 137. 

value  of,  in  zein  feeding, 
139,  140. 

Urea,  93. 

formation,  70,  71,  72,  98, 

99. 
Uric  acid,  93. 
Urine,   composition   of,   92, 

93. 


Utilization,  of  amides,  134. 
of  amino  acids,  127,  130. 
of   ammonium   salts,   133, 

134. 
of    protein,    parenterally 

introduced,  51. 

Valine,  13. 
amounts   of,  in  proteins, 
22,23. 

in  blood,  55. 
Variety  in  diet,  143. 
Vitellin,  8. 

"Wear  and  tear"  quota,  89. 

Zein,  absence  of  lysine  and 
tryptophane  in,  137. 

deficiencies  of,  in  amino 
acids,  155. 

effects  of  feeding,  141. 

feeding  experiments  with, 

139,  155. 

influence  of,  upon  growth, 
139. 

influence  of,  upon  growth 
and  maintenance,  155. 

influence  of,  upon  survi- 
val periods  of  mice,  139, 

140,  141. 


ERRATA 

Page  6,  line  21.     For  Protomines,  read  Protamines. 

Page  14,  line  5.     For  Isoleucine.  a-amino-iS-ethyl-propionic 
acid,  read  Isoleucine .  a  amino  methyl-ethyl  propionic  acid. 

Page   14,    line   25.      For  HO.C6H5.CH2.CH<cqoh, 

read  HO.C6H4.CH2.CH<co6h 

Page  16,  line  13.     For  CH2  read  CH2.COOH 

I  I 

CH2.COOH  CH2 

I  1 

CH.NH2.COOH         CH.NH2.COOH 

Page  33,  line  19.     For  1906,  read  1901. 

Page  44,  line  10.     For  CH2.  CH2.  CH2.  CH .  COOH 

I  I 

NH  NH2 


-NH2 

[2 

Arginine 

read  CH2.CH2.CH2.CH.COOH 


NH  NH2 

C^NH2 

Arginine 

Page  111,  lines  16  and  24)^      r-rrr^  ^  r^  tt  r^ 

T-,        11  o   1-        n       J -10    r  For  CeHeOs,  read  C6H12O6. 
Page  112,  Imes  7  and  18   ) 

Page  119,  line  6.      For   Ergebnisse  des   Physiologic,  read 
Ergebnisse  der  Physiologic. 

Page  121,  line  12.     For  were  metabolism,  read  were  meta- 
bolized. 


Date 

Doe 

'?Tl(/A 

DEC  I    1 

^'^b 

9Q4L 

# 

f) 

i 
] 

Cuf^^^''  '^y  AT/ON, 


